Eric J. Lang

Uniform olivocerebellar conduction time underlies Purkinje cell complex spike synchronicity in the rat cerebellum.

1. The issue of isochronicity of olivocerebellar fibre conduction time as a basis for synchronizing complex spike activity in cerebellar Purkinje cells has been addressed by latency measurement, multiple‐electrode recording and Phaseolus vulgaris leucoagglutinin (PHA‐L) tracing of climbing fibres in the adult rat. 2. The conduction time of the olivocerebellar fibres was measured by recording Purkinje cell complex spike (CS) responses from various areas of the cerebellum. The CSs were evoked by stimulating the olivocerebellar fibres near the inferior olive. In spite of a difference in length, as determined directly by light microscopy, the conduction times of different climbing fibres were quite uniform, 3.98 +/‐ 0.36 ms (mean +/‐ S.D., n = 660). 3. Multiple‐electrode recording of spontaneous Purkinje cell CS activity was employed to study the spatial extent of CS synchronicity in the cerebellar cortex. Recordings of CS were obtained from Purkinje cells located on the surface and along the walls of lobule crus 2a. The rostrocaudal band‐like distribution of simultaneous (within 1 ms) CS activity in Purkinje cells extended down the sides of the cerebellar folia to the deepest areas recorded (1.6‐2.6 mm deep). As shown in previous experiments, the distribution of simultaneous CS activity did not extend significantly (500 microns) in the mediolateral axis of the cerebellar cortex. 4. In two animals a detailed determination of the length of the olivocerebellar fibre bundles was performed by staining the fibres with PHA‐L injected into the contralateral inferior olive. This measurement included fibre bundles terminating in twenty‐six different areas, ranging from the tops of the various folia to the bottoms of the fissures in both the hemisphere and the vermis. There was a 47.5% difference between the length of the longest measured fibre bundle (15.8 mm, terminating in lobule 6b, zone A) and the length of the shortest measured fibre bundle (8.3 mm, terminating in the cortex at the base of the primary fissure, zone D), after correction for tissue shrinkage. To attain an isochronous conduction time the conduction velocities for these two fibre bundles were calculated to be 4.22 m/s and 2.37 m/s, respectively. 5. By interpolating between measured points a simple formula was derived to estimate the average length of olivocerebellar fibres terminating in any given area of the cerebellar cortex, excluding the paraflocculus, the flocculus and the most lateral regions of the hemisphere. 6. We investigated the most likely mechanisms by which conduction velocity variations with length could result in global isochronicity.(ABSTRACT TRUNCATED AT 400 WORDS)

Mnemonic correlates of unit activity in the hippocampus.

The role of the hippocampus in memory processing was examined by recording single unit activity while rats performed two different types of memory tasks. The same apparatus was used for all tasks; it consisted of two goal boxes, side by side, on the end of a runway. One goal box was white, the other was black. Experiment I used a working memory, delayed match-to-sample (DMTS) task. A trial began with a sample phase in which the rat was forced to a goal box containing a reward. The rat was then placed at the beginning of the runway again for the choice phase and allowed to enter either of the two goal boxes. Entering the goal box with the same color as that entered during the sample phase was rewarded. Experiment II used a within-subjects, within-units, design to test rats in two reference memory tasks, a cue task and a spatial task. During the cue task, the rat was rewarded for choosing the same colored goal box on each trial regardless of its spatial location. During the spatial task, the rat was rewarded for choosing the goal box in a specific location on each trial regardless of its color. During all tasks, the location of the goal boxes was changed between trials in a pseudorandom, counterbalanced fashion so that each colored goal box was on the right for half of the trials and on the left for half of the trials. During performance of the DMTS task, activity of most units was correlated with a combination of factors such as color and location, or color and phase. For example, most units showing differential activity in one of the colored goal boxes fired more when that box was in a certain spatial location, or during either the sample or choice phase. During performance of the reference memory tasks, the activity of most units was not correlated with behavior. However, the rate for some units changed between the cue and spatial tasks. When unit activity was correlated with behavior, it was dependent on a combination of dimensions such as color and spatial location. These results demonstrate that units in the hippocampus respond to combinations of stimulus dimensions such as color and spatial location, and to the temporal context necessary to solve a working memory task.

Synaptic responsiveness of interneurons of the cat lateral amygdaloid nucleus

Previous work in our laboratory has revealed that the excitability of lateral amygdaloid projection neurons is tightly regulated by GABA-mediated inhibitory postsynaptic potentials and intrinsic conductances that can be activated by synaptic inputs. Here, we studied the synaptic responsiveness of lateral amygdaloid interneurons recorded intracellularly in vivo, in the cat, to investigate their role in regulating the activity of projection cells. Interneurons were identified morphologically by their aspiny dendritic trees and physiologically by their ability to generate high frequency, non-adapting spike trains in response to depolarizing current pulses. Cortical shocks of increasing intensity generated opposite response profiles in interneurons and projection cells, with interneurons becoming progressively more excited and projection cells more inhibited. These cortically-evoked response profiles paralleled the activity of interneurons and projection cells in relation to spontaneous electroencephalographic events of differing amplitudes. Only at the lowest intensities were predominantly excitatory responses elicited in both cell types. As a result, only a narrow range of low stimulus intensities could trigger spikes in projection cells. In both cell types, the initial cortically-evoked excitatory postsynaptic potential was followed by a hyperpolarization, which was of markedly lower amplitude and duration in interneurons. In interneurons, the hyperpolarization reversed at approximately -72 mV with potassium acetate pipettes and approximately -55 mV with potassium chloride pipettes, suggesting that this inhibitory postsynaptic potential is primarily mediated by a chloride conductance. In light of previous findings indicating that inhibition in the lateral amygdaloid nucleus arises mostly from local inhibitory neurons, these results suggest that interneurons are synaptically coupled via GABAA receptors. Moreover, the opposite response profiles of interneurons and projection cells to cortical shocks indicate that interneurons play a critical role in regulating the activity of projection cells. The cellular interactions evidenced in the present study suggest that the lateral amygdaloid nucleus is endowed with an inhibitory gating mechanism that regulates information flow through the amygdala.

Serotonin Modulation of Inferior Olivary Oscillations and Synchronicity: A Multiple-electrode Study in the Rat Cerebellum

Simultaneous recording of complex spikes from multiple Purkinje cells (up to 44) in the rat cerebellum was used to examine the effects of 5‐hydroxytryptamine (serotonin, 5‐HT) on olivocerebellar function. Microinjection into the inferior olive was found to increase the average firing rate of inferior olivary neurons while slowing their oscillation frequency and increasing the coherence of their oscillations. Indeed, while the normal rostrocaudal band of synchronous activity remained unchanged, the degree of synchrony between Purkinje cell complex spikes within this band was enhanced following the 5‐HT injections. Multiple‐electrode recordings obtained from crus Ha and vermal lobule Vlb yielded qualitatively similar results; however, the effects on vermal activity were more pronounced. The effects of the 5‐HT microinjection decayed with a time course of 75 min. The half‐maximum effective concentration of 5‐HT was between 10 and 100 μM. Injections of various 5‐HT agonists and antagonists demonstrated that a 5‐HT type‐2A (5‐HT2A) receptor is the main mediator for the 5‐HT effect, which was very similar to the effect produced by injections of harmaline. However, 5‐HT and harmaline appear to have independent mechanisms since the action of harmaline was not blocked by the 5‐HT2A antagonist LY53857. A possible role for 5‐HT, as a physiological enhancer of the timing of motor function of the olivocerebellar system, is discussed.

Contextual inhibitory gating of impulse traffic in the intra-amygdaloid network.

Abstract: New data on the organization of the intra‐amygdaloid circuit is reviewed, beginning with the basolateral (BL) complex, the main input station of the amygdala for sensory afferents, and concluding with the central (CE) nucleus, an important source of projections to brain‐stem structures mediating fear responses. The BL complex is endowed with a highly divergent system of intrinsic glutamatergic connections. Yet, BL projection cells have unusually low firing rates. This apparent contradiction is explained by the presence of powerful inhibitory pressures in the BL amygdala: (1) interneurons that generate large‐amplitude inhibitory synaptic potentials and (2) projection cells that express a Ca2+‐dependent K+ current that can be activated by subthreshold synaptic inputs. Likewise, excitatory projections from the BL amygdala to the CE nucleus are controlled by clusters of GABAergic neurons, termed the intercalated (ITC) cell masses. In response to BL inputs, ITC cells generate feedforward inhibition in CE neurons. However, ITC neurons exhibit properties that allow them to modify the amount of inhibition they generate depending on the distribution of BL activity in space and time. Indeed, ITC cell masses can inhibit each other via lateromedial connections. Moreover, they express an unusual K+ conductance that modifies their response to BL inputs depending on their recent firing history. Thus, inhibitory mechanisms of the amygdala allow for flexible, context‐dependent gating of BL impulses to the CE nucleus.

Redefining the cerebellar cortex as an assembly of non-uniform Purkinje cell microcircuits

The adult mammalian cerebellar cortex is generally assumed to have a uniform cytoarchitecture. Differences in cerebellar function are thought to arise primarily through distinct patterns of input and output connectivity rather than as a result of variations in cortical microcircuitry. However, evidence from anatomical, physiological and genetic studies is increasingly challenging this orthodoxy, and there are now various lines of evidence indicating that the cerebellar cortex is not uniform. Here, we develop the hypothesis that regional differences in properties of cerebellar cortical microcircuits lead to important differences in information processing.

Block of Inferior Olive Gap Junctional Coupling Decreases Purkinje Cell Complex Spike Synchrony and Rhythmicity

Inferior olivary (IO) neurons are electrotonically coupled by gap junctions. This coupling is thought to underlie synchronous complex spike (CS) activity generated by the olivocerebellar system in Purkinje cells, and also has been hypothesized to be necessary for IO neurons to generate spontaneous oscillatory activity. These characteristics of olivocerebellar activity have been proposed to be central to the role of this system in motor coordination. However, the relationship of gap junction coupling between IO neurons to synchronous and rhythmic CS activity has never been directly tested. Thus, to address this issue, multiple electrode recordings were obtained from crus 2a Purkinje cells, and carbenoxolone, a gap junction blocker, was injected into the IO. Carbenoxolone reduced CS synchrony by 50% overall, but in some experiments, >80% reductions were achieved. Carbenoxolone also reduced the average firing rate by 50%, suggesting that electrical coupling is a significant source of excitation for IO neurons. Moreover, carbenoxolone caused a reduction in the ∼10 Hz rhythmicity of CS activity, and this reduction was correlated with the extent to which the injection reduced CS synchrony. Lastly, carbenoxolone was found to reverse or prevent changes in synchrony that are normally induced by injection of GABAA and glutamate receptor antagonists into the IO, suggesting that the effects of these drugs on CS synchrony patterns require electrical coupling of IO neurons. In sum, our results provide direct evidence that electrical coupling of IO neurons underlies synchronous CS activity, and suggest important roles for this coupling in shaping other aspects of IO spiking patterns.

Olivocerebellar modulation of motor cortex ability to generate vibrissal movements in rat

The vibrissal movements known as whisking are generated in a pulsatile, or non‐continuous, fashion and comprise sequences of brief regularly spaced movements. These rhythmic timing sequences imply the existence of periodically issued motor commands. As inferior olivary (IO) neurones generate periodic synchronous discharges that could provide the underlying timing signal, this possibility was tested by determining whether the olivocerebellar system modulates motor cortex (MCtx)‐triggered whisker movements in rats. Trains of current pulses were applied to MCtx, and the resulting whisker movements were recorded using a high speed video camera. The evoked movement patterns demonstrated properties consistent with the existence of an oscillatory motor driving rhythm. In particular, movement amplitude showed a bell‐shaped dependence on stimulus frequency, with a peak at 11.5 ± 2.3 Hz. Moreover, movement trajectories showed harmonic and subharmonic entrainment patterns within specific stimulus frequency ranges. By contrast, movements evoked by facial nerve stimulation showed no such frequency‐dependent properties. To test whether the IO was the oscillator in question, IO neuronal properties were modified in vivo by intra‐IO picrotoxin injection, which enhances synchronous oscillatory IO activity and reduces its natural frequency. The ensuing changes in the evoked whisker patterns were consistent with these pharmacological effects. Furthermore, in cerebellectomized rats, oscillatory modulation of MCtx‐evoked movements was greatly reduced, and intra‐IO picrotoxin injections did not affect the evoked movement patterns. Additionally, multielectrode recording of Purkinje cell complex spikes showed a temporal correlation of olivocerebellar activity during MCtx stimulus trains to evoked movement patterns. In sum, the results indicate that MCtxs ability to generate movements is modulated by an oscillatory signal arising in the olivocerebellar system.

Spontaneous activity of the perirhinal cortex in behaving cats

The perirhinal cortex lies at the interface between the neocortex and allocortex. Whether the perirhinal cortex expresses spontaneous electroencephalographic rhythms that are characteristic of the allocortex and/or of the neocortex is unknown. Thus, the present investigation was undertaken to characterize the activity of the perirhinal cortex with respect to various electroencephalographic rhythms that are displayed by neocortical areas or the entorhino-hippocampal system during different behavioral states of vigilance in chronically-implanted cats. Although perirhinal and neocortical electroencephalograms underwent similar state-dependent changes in amplitude, the ubiquitous neocortical sleep spindles were absent from the perirhinal cortex. In addition, while the slow sleep oscillation (0.5-1 Hz), which is pervasive in the neocortex, was present in the perirhinal cortex, its temporal relation to the neocortical oscillation was highly variable. In contrast, a high degree of correlation was found between perirhinal and entorhinal electroencephalographic activities in all behavioral states. In particular, during waking and paradoxical sleep, multiple simultaneously recorded entorhinal and perirhinal sites displayed an oscillation in the theta range which was highly correlated. To rule out the possibility that the perirhinal theta oscillation reflected volume conduction from neighboring structures, single-unit recordings were performed. Spike-triggered averages and peri-event histograms revealed that perirhinal cells displayed a statistically significant theta-related modulation of their spontaneous activity, albeit weaker than that observed in the entorhinal cortex. Thus, from the standpoint of spontaneous electroencephalographic rhythms, the perirhinal cortex is more closely related to the entorhino-hippocampal system than to the neocortex.

High-Resolution Measurement of the Time Course of Calcium-Concentration Microdomains at Squid Presynaptic Terminals

Transmitter release is considered to be a secretory event triggered by localized calcium influx which, by binding to a low-affinity Ca2+ site at the presynaptic active zone, initiates vesicular exocytosis (1-7). In previous experiments with aequorin-loaded presynaptic terminals we visualized, upon tetanic presynaptic stimulation, small points of light produced by calcium concentration microdomains of about 300 microM (5). These microdomains had a diameter of about 0.5 microns (5) and covered 5-10% of the total presynaptic membrane with an average density of 8.4 microns2 per 100 microns2, corresponding closely to the size and distribution of the active zones in that junction (6, 7). To understand in more detail the nature of these concentration microdomains, we obtained rapid video images (400/s) after injecting the photoprotein n-aequorin-J into the presynaptic terminals of squid giant synapses. Using that experimental approach, we determined that microdomains evoked by presynaptic spike activation had a duration of about 800 microseconds. Spontaneous quantum emission domains (QEDs) observed at about the same locations as the microdomains were smaller in amplitude, shorter in duration, and less frequent. These results illustrate the time course of the calcium concentration profiles responsible for transmitter release. Their extremely short duration compares closely with that of calcium current flow during a presynaptic action potential and indicates that, as theorized in the past (6-8), intracellular calcium concentration at the active zone remains high only for the duration of transmembrane calcium flow.