Integration of broadband conductance input in rat somatosensory cortical inhibitory interneurons: an inhibition-controlled switch between intrinsic and input-driven spiking in fast-spiking cells.

Quantitative understanding of the dynamics of particular cell types when responding to complex, natural inputs is an important prerequisite for understanding the operation of the cortical network. Different types of inhibitory neurons are connected by electrical synapses to nearby neurons of the same type, enabling the formation of synchronized assemblies of neurons with distinct dynamical behaviors. Under what conditions is spike timing in such cells determined by their intrinsic dynamics and when is it driven by the timing of external input? In this study, we have addressed this question using a systematic approach to characterizing the input-output relationships of three types of cortical interneurons (fast spiking [FS], low-threshold spiking [LTS], and nonpyramidal regular-spiking [NPRS] cells) in the rat somatosensory cortex, during fluctuating conductance input designed to mimic natural complex activity. We measured the shape of average conductance input trajectories preceding spikes and fitted a two-component linear model of neuronal responses, which included an autoregressive term from its own output, to gain insight into the input-output relationships of neurons. This clearly separated the contributions of stimulus and discharge history, in a cell-type dependent manner. Unlike LTS and NPRS cells, FS cells showed a remarkable switch in dynamics, from intrinsically driven spike timing to input-fluctuation-controlled spike timing, with the addition of even a small amount of inhibitory conductance. Such a switch could play a pivotal role in the function of FS cells in organizing coherent gamma oscillations in the local cortical network. Using both pharmacological perturbations and modeling, we show how this property is a consequence of the particular complement of voltage-dependent conductances in these cells.

Quantifying noise-induced stability of a cortical fast-spiking cell model with Kv3-channel-like current.

Population oscillations in neural activity in the gamma (>30 Hz) and higher frequency ranges are found over wide areas of the mammalian cortex. Recently, in the somatosensory cortex, the details of neural connections formed by several types of GABAergic interneurons have become apparent, and they are believed to play a significant role in generating these oscillations through synaptic and gap-junctional interactions. However, little is known about the mechanism of how such oscillations are maintained stably by particular interneurons and by their local networks, in a noisy environment with abundant synaptic inputs. To obtain more insight into this, we studied a fast-spiking (FS)-cell model including Kv3-channel-like current, which is a distinctive feature of these cells, from the viewpoint of nonlinear dynamical systems. To examine the specific role of the Kv3-channel in determining oscillation properties, we analyzed basic properties of the FS-cell model, such as the bifurcation structure and phase resetting curves (PRCs). Furthermore, to quantitatively characterize the oscillation stability under noisy fluctuations mimicking small fast synaptic inputs, we applied a recently developed method from random dynamical system theory to estimate Lyapunov exponents, both for the original four-dimensional dynamics and for a reduced one-dimensional phase-equation on the circle. The results indicated that the presence of the Kv3-channel-like current helps to regulate the stability of noisy neural oscillations and a transient-period length to stochastic attractors.

Phase resetting curves and oscillatory stability in interneurons of rat somatosensory cortex.

Synchronous oscillations in neural activity are found over wide areas of the cortex. Specific populations of interneurons are believed to play a significant role in generating these synchronized oscillations through mutual synaptic and gap-junctional interactions. Little is known, though, about the mechanism of how oscillations are maintained stably by particular types of interneurons and by their local networks. To obtain more insight into this, we measured membrane-potential responses to small current-pulse perturbations during regular firing, to construct phase resetting curves (PRCs) for three types of interneurons: nonpyramidal regular-spiking (NPRS), low-threshold spiking (LTS), and fast-spiking (FS) cells. Within each cell type, both monophasic and biphasic PRCs were observed, but the proportions and sensitivities to perturbation amplitude were clearly correlated to cell type. We then analyzed the experimentally measured PRCs to predict oscillation stability, or firing reliability, of cells for a complex stochastic input, as occurs in vivo. To do this, we used a method from random dynamical system theory to estimate Lyapunov exponents of the simplified phase model on the circle. The results indicated that LTS and NPRS cells have greater oscillatory stability (are more reliably entrained) in small noisy inputs than FS cells, which is consistent with their distinct types of threshold dynamics.

Rate coding and spike-time variability in cortical neurons with two types of threshold dynamics.

Neurons and dynamical models of spike generation display two different classes of threshold behavior: type 1 [firing frequency vs. current (f-I) relationship is continuous at threshold] and type 2 (discontinuous f-I). With steady current or conductance stimulation, regular-spiking (RS) pyramidal neurons and fast-spiking (FS) inhibitory interneurons in layer 2/3 of somatosensory cortex exhibit type 1 and type 2 threshold behaviors, respectively. We compared the postsynaptic firing variability of type 1 RS and type 2 FS cells, during naturalistic, fluctuating conductance input. In RS neurons, increasing the level of independently random, shunting inhibition caused a monotonic increase in spike reliability, whereas in FS interneurons, there was an optimum level of shunting inhibition for achieving the most reliable spike generation and the most precise spike-time encoding. This was observed over a range of different degrees of synchrony, or correlation, in the input. RS cells displayed a progressive rise in spike jitter during natural-like transient burst inputs, whereas for FS cells, jitter was mostly kept low. Furthermore, RS cells showed encoding of the input level in the spike shape, whereas FS cells did not. These differences between the two cell types are consistent with a role of RS neurons as rate-coding integrators, and a role of FS neurons as resonators controlling the coherence of synchronous firing.

Spatio-temporal cholinergic modulation in cultured networks of rat cortical neurons: spontaneous activity.

Activation of the cholinergic innervation of the cortex has been implicated in sensory processing, learning, and memory. At the cellular level, acetylcholine both increases excitability and depresses synaptic transmission, and its effects on network firing are hard to predict. We studied the effects of carbachol, a cholinergic agonist, on network firing in cultures of rat cortical neurons, using electrode arrays to monitor the activity of large numbers of neurons simultaneously. These cultures show stable spontaneous synchronized burst firing which propagates through dense synaptic connections. Carbachol (10-50 microM), acting through muscarinic receptors, was found to induce a switch to asynchronous single-spike firing and to result in a loss of regularity and fragmentation of the burst structure. To obtain a quantitative measure of cholinergic actions on cortical networks, we applied a cluster Poisson-process model to sets of paralleled spike-trains in the presence and absence of carbachol. This revealed that the time series can be well-characterized by such a simple model, consistent with the observed 1/f(b)-like spectra (0.04

Spatio-temporal cholinergic modulation in cultured networks of rat cortical neurons: evoked activity.

We studied the effects of carbachol, a cholinergic agonist, on extracellularly evoked firing of networks in mature cultures of rat cortical neurons, using multi-electrode arrays to monitor the activity of large numbers of neurons simultaneously. These cultures show evoked burst firing which propagates through dense synaptic connections. When a brief voltage pulse was applied to one extracellular electrode, spiking electrical responses were evoked in neurons throughout the network. The response had two components: an early phase, terminating within 30-80 ms, and a late phase which could last several hundreds of milliseconds. Action potentials evoked during the early phase were precisely timed, with only small jitter. In contrast, the late phase characteristically showed clusters of electrical activity with significant spatio-temporal fluctuations. The late phase was suppressed by applying a relatively small amount of carbachol (5 microM) in the external solution, even though the spontaneous firing rate was not significantly changed. Carbachol increased both the spike-timing precision and the speed of propagation of population spikes, and selectively increased the firing coincidence in a subset of neuron pairs in the network, while suppressing late variable firing in responses. Hence, the results give quantitative support for the idea that cholinergic activation in the cortex has a general role of focusing or enhancing significant associative firing of neurons.

The interpretation of current-clamp recordings in the cell-attached patch-clamp configuration.

In these experiments we have investigated the feasibility and accuracy of recording steady-state and dynamic changes in transmembrane potential noninvasively across an intact cell-attached patch using the current-clamp mode of a conventional patch-clamp amplifier. Using an equivalent circuit mimicking simultaneous whole-cell voltage-clamp and cell-attached current-clamp recordings we have defined both mathematically and experimentally the relationship between the membrane patch resistance, the seal resistance, and the fraction of the whole-cell potential recorded across an intact membrane patch. This analysis revealed a steep increase in the accuracy of recording of steady-state membrane potential as the seal/membrane ratio increases from 0. The recording accuracy approaches 100% as the seal/membrane ratio approaches infinity. Membrane potential measurements across intact cell-attached patches in rat basophilic leukemia cells and rat megakaryocytes revealed a surprisingly high degree of accuracy and demonstrated the ability of this noninvasive technique to follow dynamic changes in potential in nonexcitable cells.

Threshold firing frequency-current relationships of neurons in rat somatosensory cortex: type 1 and type 2 dynamics.

Neurons and dynamical models of spike generation display two different types of threshold behavior, with steady current stimulation: type 1 [the firing frequency vs. current (f-I) relationship is continuous at threshold) and type 2 (discontinuous f-I)]. The dynamics at threshold can have profound effects on the encoding of input as spikes, the sensitivity of spike generation to input noise, and the coherence of population firing. We have examined the f-I and frequency-conductance (f-g) relationships of cells in layer 2/3 of slices of young (15-21 DIV) rat somatosensory cortex, focusing in detail on the nature of the threshold. Using white-noise stimulation, we also measured firing frequency and interspike interval variability as a function of noise amplitude. Regular-spiking (RS) pyramidal neurons show a type 1 threshold, consistent with their well-known ability to fire regularly at very low frequencies. In fast-spiking (FS) inhibitory interneurons, although regular firing is supported over a wide range of frequencies, there is a clear discontinuity in their f-I relationship at threshold (type 2), which has not previously been highlighted. FS neurons are unable to support maintained periodic firing below a critical frequency fc, in the range of 10 to 30 Hz. Very close to threshold, FS cells switch irregularly between bursts of periodic firing and subthreshold oscillations. These characteristics mean that the dynamics of RS neurons are well suited to encoding inputs into low-frequency firing rates, whereas the dynamics of FS neurons are suited to maintaining and quickly synchronizing to gamma and higher-frequency input.

Estimation of fetal weight by ultrasound prior to 33 weeks gestation.

The aim of this study was to develop an accurate formula for the ultrasonic prediction of fetal weight for infants < 33 weeks gestational age and < or = 1500 g birthweight. The subjects comprised live births free of lethal malformations or chromosomal anomalies, < 33 weeks gestational age and with birthweights +/- 1500 g born in the Royal Women's Hospital between January 1990 and March 1996. All subjects had accurate gestational age confirmed by ultrasound prior to 20 weeks gestation and ultrasound measurements within 72 hours of birth of biparietal diameter (BPD), femur length (FL) and abdominal circumference (AC). A formula with the highest explained variance was computed by linear regression analysis using the three fetal variables in various combinations from 54 infants born between January 1990 and December 1993. The optimal formula was: Log(10)birthweight = 0.714627 + 0.077362.AC + 0.058758.BPD + 0.287037.FL - 0.011274.AC.FL. The new formula was more accurate compared with existing formulae when tested in a separate cohort of 39 infants born between January 1994 and March 1996.

Postsynaptic variability of firing in rat cortical neurons: the roles of input synchronization and synaptic NMDA receptor conductance.

Neurons in the functioning cortex fire erratically, with highly variable intervals between spikes. How much irregularity comes from the process of postsynaptic integration and how much from fluctuations in synaptic input? We have addressed these questions by recording the firing of neurons in slices of rat visual cortex in which synaptic receptors are blocked pharmacologically, while injecting controlled trains of unitary conductance transients, to electrically mimic natural synaptic input. Stimulation with a Poisson train of fast excitatory (AMPA-type) conductance transients, to simulate independent inputs, produced much less variability than encountered in vivo. Addition of NMDA-type conductance to each unitary event regularized the firing but lowered the precision and reliability of spikes in repeated responses. Independent Poisson trains of GABA-type conductance transients (reversing at the resting potential), which simulated independent activity in a population of presynaptic inhibitory neurons, failed to increase timing variability substantially but increased the precision of responses. However, introduction of synchrony, or correlations, in the excitatory input, according to a nonstationary Poisson model, dramatically raised timing variability to in vivo levels. The NMDA phase of compound AMPA-NMDA events conferred a time-dependent postsynaptic variability, whereby the reliability and precision of spikes degraded rapidly over the 100 msec after the start of a synchronous input burst. We conclude that postsynaptic mechanisms add significant variability to cortical responses but that substantial synchrony of inputs is necessary to explain in vivo variability. We suggest that NMDA receptors help to implement a switch from precise firing to random firing during responses to concerted inputs.

Propagation of spontaneous synchronized activity in cortical slice cultures recorded by planar electrode arrays.

The spatial propagation of synchronized activity in cortical slice cultures was characterized by multi-site extracellular recording. Spontaneous activity was studied in normal culture medium, and in bicuculline- or kainic acid-containing media. A common feature in all these conditions was that activity was generated first in superficial layers (i.e., layer I/II) before spreading over the whole area of the slice. In culture medium or bicuculline-containing medium, the initiation site of the activity was not constant and showed a large variety of patterns of horizontal propagation. Kainic acid induced epileptiform activity, consisting of intense initial bursts followed by repetitive after-discharges. Though the patterns of spatial propagation of the bursts were variable as in the other conditions, the after-discharges followed a constant path. Cross-correlation analysis indicated that the network moved in a graded fashion to a steady state during the sequence of after-discharges.

A new class of neurotoxin from wasp venom slows inactivation of sodium current.

The effects of alpha-pompilidotoxin (alpha-PMTX), a new neurotoxin isolated from the venom of a solitary wasp, were studied on the neuromuscular synapses in lobster walking leg and the rat trigeminal ganglion (TG) neurons. Paired intracellular recordings from the presynaptic axon terminals and the innervating lobster leg muscles revealed that alpha-PMTX induced long bursts of action potentials in the presynaptic axon, which resulted in facilitated excitatory and inhibitory synaptic transmission. The action of alpha-PMTX was distinct from that of other known facilitatory presynaptic toxins, including sea anemone toxins and alpha-scorpion toxins, which modify the fast inactivation of Na+ current. We further characterized the action of alpha-PMTX on Na+ channels by whole-cell recordings from rat trigeminal neurons. We found that alpha-PMTX slowed the Na+ channels inactivation process without changing the peak current-voltage relationship or the activation time course of tetrodotoxin (TTX)-sensitive Na+ currents, and that alpha-PMTX had voltage-dependent effects on the rate of recovery from Na+ current inactivation and deactivating tail currents. The results suggest that alpha-PMTX slows or blocks conformational changes required for fast inactivation of the Na+ channels on the extracellular surface. The simple structure of alpha-PMTX, consisting of 13 amino acids, would be advantageous for understanding the functional architecture of Na+ channel protein.

Determining the activation time course of synaptic AMPA receptors from openings of colocalized NMDA receptors.

Excitatory postsynaptic currents (EPSCs) in most mammalian central neurons have a fast alpha-amino-3-hydroxy-5-methyl-4-isoazole-proprionic acid (AMPA) receptor-mediated component, lasting a few milliseconds, and a slow N-methyl-D-aspartic acid (NMDA)-receptor-mediated component, lasting hundreds of milliseconds. The time course of the AMPA phase is crucial in the integrative function of neurons, but measuring it accurately is often confounded by cable filtering between the recording electrode and the synapse. We describe a method for recovering the AMPA phase of individual EPSCs by determining the impulse response of the cable filter from single NMDA channel transitions in the slow tails of the same EPSC, then deconvolving the measured AMPA current. Using simulations, we show that filtering of an AMPA conductance transient in a voltage-clamped dendrite behaves in an almost perfectly linear fashion. Expressions are derived for the time course of single channel transitions and the AMPA phase filtered through a voltage-clamped cable or a single exponential filter, using a kinetic model for AMPA receptor activation. Fitting these expressions to experimental records directly estimates the underlying kinetics of the AMPA phase. Example measurements of spontaneous EPSCs in cultured nonpyramidal rat cortical neurons yielded rising time constants of 0.2-0.8 ms, and decay time constants of 1.3-2 ms at 23-25 degrees C.

Simultaneous induction of pathway-specific potentiation and depression in networks of cortical neurons.

Activity-dependent modification of synaptic efficacy is widely recognized as a cellular basis of learning, memory, and developmental plasticity. Little is known, however, of the consequences of such modification on network activity. Using electrode arrays, we examined how a single, localized tetanic stimulus affects the firing of up to 72 neurons recorded simultaneously in cultured networks of cortical neurons, in response to activation through 64 different test stimulus pathways. The same tetanus produced potentiated transmission in some stimulus pathways and depressed transmission in others. Unexpectedly, responses were homogeneous: for any one stimulus pathway, neuronal responses were either all enhanced or all depressed. Cross-correlation of responses with the responses elicited through the tetanized site revealed that both enhanced and depressed responses followed a common principle: activity that was closely correlated before tetanus with spikes elicited through the tetanized pathway was enhanced, whereas activity outside a 40-ms time window of correlation to tetanic pathway spikes was depressed. Response homogeneity could result from pathway-specific recurrently excitatory circuits, whose gain is increased or decreased by the tetanus, according to its cross-correlation with the tetanized pathway response. The results show how spatial responses following localized tetanic stimuli, although complex, can be accounted for by a simple rule for activity-dependent modification.

Strengthening of synchronized activity by tetanic stimulation in cortical cultures: application of planar electrode arrays.

Rat cortical neurons were cultured on planar electrode arrays with 64 embedded electrodes. Whole-cell recording from single neurons and multisite extracellular recording were carried out simultaneously in the cultured cortical networks, and the effects of focal tetanic stimulation of the culture were studied. Both the number of action potentials and the propagation velocity of stimulated bursts were increased after tetanic stimulation. These changes were associated with a marked increase in the number of late components in the synaptic current, but with little or no increase in the early peak synaptic current. The effects of tetanic stimulation were consistent with a widespread increase in the reliability of monosynaptic transmission.

Modification of parallel activity elicited by propagating bursts in developing networks of rat cortical neurones.

Networks of cultured cortical neurones exhibit regular, synchronized, propagating bursts which are synaptically mediated, and which are hypothesized to play a part in activity-dependent formation of connections during development in vivo. The relationship between the strength of synaptic connections and the characteristics of synchronized propagating bursting, however, is unclear. Modification of synchronized activity in cortical cultures in response to electrical stimulation was examined using multisite electrode array recording. By measuring the response of the network to weak, localized, test stimulation (TS), we observed a potentiation of activity following a relatively stronger inducing stimulation (IS). This potentiation was evident as an increased probability of eliciting bursts by TS, an increased frequency of spontaneous bursts and number of spikes per burst, and increased speed of burst propagation, and it lasted for at least 20 min. Changing the parameters of IS revealed that high frequency tetanic stimulation is not necessary to induce potentiation, while it is essential for IS to produce a regeneratively propagating burst. The results provide a direct demonstration of modification of both the spatial and temporal characteristics of synchronized network activity, and suggest an important physiological role for propagating synchronized bursting, as a mechanism for inducing plastic modifications in the developing cortex.

Intracellular inositol 1,3,4,5-tetrakisphosphate enhances the calcium current in hippocampal CA1 neurones of the gerbil after ischaemia.

1. To examine the role of the phosphoinositide cascade triggered by disturbed Ca2+ homeostasis in ischaemic neurones, inositol 1,3,4,5-tetrakisphosphate (InsP4) was applied to the cytoplasmic face of membrane patches isolated from CA1 pyramidal neurones in the gerbil hippocampus. 2. In outside-out recordings, InsP4 induced an inward current which was increased by raising the extracellular [Ca2+]. In contrast, no clear channel openings could be observed in patches from neurones of sham-operated gerbils. 3. Open probabilities of InsP4-activated channels were significantly decreased upon application of omega-conotoxin but were not affected by omega-agatoxin or nifedipine. 4. In inside-out patches using high concentrations of Ca2+, Ba2+ or Sr2+ in the pipette solution, InsP4 enhanced inward currents. 5. Application of the isomers of InsP4 slightly enhanced the currents, but inositol 1,4,5-trisphosphate (InsP3) had no effect. 6. In the absence of InsP4 there was a single main Ba2+ current peak of 4.0 pA in amplitude, whereas upon its application two main peaks of 3.0 and 7.2 pA were present. 7. The open probabilities of these channels were apparently increased by InsP4. 8. These findings support the view that a disturbed phosphoinositide cascade occurs in the hippocampal pyramidal neurones after ischaemia and the InsP4 thus formed plays an important role in promoting the Ca2+ accumulation which results in neuronal death.

Correlation between nuchal thickness and abnormal karyotype in first trimester fetuses.

OBJECTIVE: To evaluate the accuracy of ultrasound measurement of nuchal thickness in first trimester fetuses for predicting fetal karyotype. DESIGN: A prospective study of the nuchal thickness of fetuses measured during an ultrasound examination in all women undergoing first trimester chorionic villus sampling (CVS). SETTING: Two major public hospitals and two associated private practices between 7 September 1993 and 6 September 1994. PARTICIPANTS: Pregnant women with various indications for CVS (in 82% because of maternal age). RESULTS: 1306 women underwent CVS, including 11 with twin pregnancies: 1317 fetuses were tested. Karyotype results were obtained for 1312 fetuses: 41 (3.1%) had an abnormal karyotype, and 20 of these (49%) had a nuchal thickness measurement of 3 mm or more, compared with 44 (3.5%) of the 1271 fetuses with a normal karyotype. Of the 21 fetuses shown to have trisomy 21, 12 would have been detected if a nuchal thickness of 3 mm or more had been used as an indicator, giving a sensitivity of 57%. Nuchal thickness measurements of 1 or 2 mm excluded trisomy 21 with a negative predictive value of 99.3%. Fetuses with moderate nuchal thickening, normal karyotype and no other problems noted on the initial ultrasound scan had neonatal outcomes similar to those in the general obstetric population. CONCLUSION: Nuchal thickening in the first trimester (10 weeks on) of pregnancy in a high risk population is a powerful indicator of increased risk of aneuploidy.

Spontaneous periodic synchronized bursting during formation of mature patterns of connections in cortical cultures.

Long-term recording of spontaneous activity in cultured cortical neuronal networks was carried out using substrates containing multi-electrode arrays. Spontaneous uncorrelated firing appeared within the first 3 days and transformed progressively into synchronized bursting within a week. By 30 days from the establishment of the culture, the network exhibited a complicated non-periodic, synchronized activity pattern which showed no changes for more than 2 months and thus represented the mature state of the network. Pharmacological inhibition of activity only during the period when regular synchronized bursting was observed was capable of producing a different mature activity pattern from the control. These results suggest that periodic synchronized bursting plays a critical role in the development of synaptic connections.

The mechanisms of generation and propagation of synchronized bursting in developing networks of cortical neurons.

The characteristics and mechanisms of synchronized firing in developing networks of cultured cortical neurons were studied using multisite recording through planar electrode arrays (PEAs). With maturation of the network (from 3 to 40 d after plating), the frequency and propagation velocity of bursts increased markedly (approximately from 0.01 to 0.5 Hz and from 5 to 100 mm/sec, respectively), and the sensitivity to extracellular magnesium concentration (0-10 mM) decreased. The source of spontaneous bursts, estimated from the relative delay of onset of activity between electrodes, varied randomly with each burst. Physical separation of synchronously bursting networks into several parts using an ultraviolet laser, divided synchronous bursting into different frequencies and phases in each part. Focal stimulation through the PEA was effective at multiple sites in eliciting bursts, which propagated over the network from the site of stimulation. Stimulated bursts exhibited both an absolute refractory period and a relative refractory period, in which partially propagating bursts could be elicited. Periodic electrical stimulation (at 1 to 30 sec intervals) produced slower propagation velocities and smaller numbers of spikes per burst at shorter stimulation intervals. These results suggest that the generation and propagation of spontaneous synchronous bursts in cultured cortical neurons is governed by the level of spontaneous presynaptic firing, by the degree of connectivity of the network, and by a distributed balance between excitation and recovery processes.