Cerebello-cerebellar responses mediated via climbing fibres

Summary A sequence of three evoked potentials was recorded on the paramedian lobule of the cat cerebellum following single shock stimulation of the intermediate portion of the ipsilateral anterior lobe. The second and third responses have been analysed using micro-electrode recording techniques and each was shown to reflect near-synchronous climbing fibre activation of many Purkinje cells in the paramedian lobule. The first response has not been fully studied but did not involve the climbing fibres.The first climbing fibre evoked potential was constant in amplitude and latency and able to follow at stimulation frequencies as high as 350/sec. It was scarcely affected by section of the ipsilateral cerebellar peduncles. The second was very variable in amplitude, never followed at frequencies greater than c. 10/sec and was completely abolished by pedunculotomy. Intracellular recordings from Purkinje cells showed that the first climbing fibre response was associated with a single excitatory post-synaptic potential but the second with excitatory post-synaptic potentials which were usually multiple.The above characteristics, together with the results of impulse collision experiments, have been satisfactorily accounted for by postulating that the climbing fibre afferents branch widely after entering the cerebellum. The earlier response arose as a result of axon reflexes and the later response as a result of orthodromic impulses generated by the cells of origin of the fibres. Evidence is presented which suggests that some at least of these cells were situated in the inferior olive.     

Interaction experiments on the responses evoked in Purkinje cells by climbing fibres

1. The uniquely powerful excitatory synaptic action of a single climbing fibre on a Purkinje cell in the cerebellum of the cat was tested during the intense and prolonged inhibitory action produced by the parallel fibre, basket and stellate cell system. There was depression of the later spike discharges, but the initial discharge was never suppressed.2. With intracellular recording the excitatory post-synaptic potential was depressed during the initial phase (about 10 msec) of the inhibitory action, but there was a later increase with a time course resembling the latter part of the inhibitory hyperpolarization. An explanation of these and other effects is given in terms of conventional ideas of excitatory and inhibitory synaptic interaction.3. These observations on single Purkinje cells, particularly with intracellular recording, have helped in formulating a provisional explanation of the finding that during inhibition there is an increase in the negative field potential evoked by a climbing fibre volley.4. The excitatory action of a climbing fibre synapse is shown to be greatly depressed immediately after a preceding activation and recovery takes hundreds of milliseconds. By the collision technique it is shown that the same climbing fibre is activated by inferior olive and juxta-fastigial stimulation.5. With rapid repetitive activation there was initially a progressive decline in the effectiveness of each successive impulse, but a steady level was soon reached. On cessation of a tetanus of twenty or more impulses there was a delayed recovery of the depolarization, which suggests a continued action of the accumulated transmitter.6. With extracellular recording repetitive spike initiation continued with stimulation frequencies as high as 100/sec, but at still higher frequencies spikes were depressed by the intense synaptically evoked depolarization. On cessation of the stimulation after-discharge often developed as the depolarization declined. The prolonged after-discharges following severe tetani suggest that there is a very effective accumulation of the transmitter.     

Distribution of climbing fibres on cerebellar Purkinje cells in X-irradiated rats. An electrophysiological study.

1. The distribution of climbing fibres on cerebellar Purkinje cells has been studied with intracellular recordings in X-irradiated and normal rats. 2. In the treated rats, multiple steps in the post-synaptic potential were elicited in 57% of the Purkinje cells by graded stimulation of the climbing fibres, the response was all-or-none in character in the other cells and in all Purkinje cells recorded in normal animals. In the neurones exhibiting the former type of response, no collision was seen along the afferent fibres during interaction experiments between just-threshold juxtafastigial and maximal olivary stimulations, whereas a collision always occurred when all-or-none responses were recorded. 3. These results show that in X-irradiated rats, the majority of Purkinje cells have a multiple innervation by two to four climbing fibres, instead of the one-to-one relationship seen normally. 4. Input resistances and total electrotonic lengths of Purkinje cells were measured in normal and treated rats. Mean values for these two parameters were higher than normal in multiply innervated cells. 5. Mean time course and mean current for reversal of the post-synaptic potential elicited in Purkinje cells by stimulation of the climbing fibres were nearly the same in mono- and in multiply innervated neurones. In multiply innervated cells, time courses and currents for reversal were independent of the size of the response or varied slightly with it, suggesting that the climbing fibres involved innervated territories whose electrotonic distance from the recording site were either the same or slightly different. 6. Interactions between two all-or-none steps of the graded post-synaptic potential evoked in multiply innervated cells by juxtafastigial and olivary stimulations revealed either a very weak or a very marked shunting effect between synapses of the two climbing fibres involved. 7. These results indicate that the over-all distribution of climbing fibre synapses on multiply innervated Purkinje cells is not grossly abnormal and that two fibres contacting a given cell can be either intermingled on the same dendrites, or segregated on distinct dendritic branches. 8. In general, the present study does not suggest the existence of a strong competition among climbing fibres innervating each Purkinje cell during development at least when granule cells are absent.     

Inhibition of the inferior olive during conditioned responses in the decerebrate ferret

Output from the interpositus nucleus can inhibit the inferior olive, probably via the GABA-ergic nucleo-olivary pathway. It has been suggested that the function of this inhibition might be to regulate synaptic plasticity resulting from parallel fibre/climbing fibre interaction in cerebellar Purkinje cells, by providing negative feedback information to the olive. Thus, when a learned response, generated by the interpositus nucleus, reaches a sufficient amplitude, the olive would be inhibited and further learning blocked. This suggestion was tested in a classical conditioning paradigm. Decerebrate ferrets were trained using electrical skin stimulation of the forelimb as the conditioned stimulus (CS) and periorbital stimulation as the unconditioned stimulus (US). Climbing fibre responses evoked in Purkinje cells by the US were recorded as surface field potentials in the part of the c3 zone controlling eyeblink. It was found that the CS did not inhibit the olive at the beginning of training, but when conditioned responses were large, the olive was inhibited by the CS in some animals. After a number of unpaired CS presentations, which caused extinction of the conditioned response, the inhibition disappeared. The size of individual conditioned responses correlated negatively with the size of the climbing fibre responses evoked by the US. Climbing fibre responses evoked by direct stimulation of the olive were also inhibited. It was concluded that cerebellar output during performance of a conditioned response inhibits the inferior olive. The results thus support the hypothesis of a cerebellar locus of conditioning and are consistent with the proposed role of cerebello-olivary inhibition.     

Mode of induction of long-term depression at parallel fibre--Purkinje cell synapses in rabbit cerebellar cortex

In a superficial folium of the dorsal paraflocculus of high decerebrate rabbits, extracellular unitary spikes were recorded from a Purkinje cell, while two parallel fibre beams impinging onto that Purkinje cell were separately stimulated in the molecular layer. Climbing fibre afferents were stimulated at the contralateral inferior olive. Quisqualate was ionophoretically applied to the dendrite of the Purkinje cell intersecting one of the stimulated parallel fibre beams (test beam). Long-term depression (longer than 45 min) occurred in Purkinje cell responsiveness to the test beam, but not to the other beam (control beam), when quisqualate was applied for 4 min in conjunction with 2 Hz stimulation of climbing fibres. This effect was completely abolished by simultaneous application of a glutamate blocker, kynurenate, during conjunctive quisqualate-climbing fibre stimulation. Application of quisqualate alone caused a small degree of depression in parallel fibre-Purkinje cell transmission. This effect was abolished when spontaneous activity of climbing fibres was blocked by injection of tetrodotoxin or lidocaine to the contralateral inferior olive, and therefore was due to conjunction of quisqualate with spontaneous climbing fibre inputs that normally occurred at 0.5-1.2 Hz. These findings suggest that the occurrence of long-term depression is strictly dependent on conjunction of climbing fibre activity with quisqualate receptor activation.     

Principal neuron spiking: neither necessary nor sufficient for cerebral blood flow in rat cerebellum

Neuronal activity, cerebral blood flow, and metabolic responses are all strongly coupled, although the mechanisms behind the coupling remain unclear. One of the key questions is whether or not increases in spiking activity in the stimulated neurons are sufficient to drive the activity-dependent rises in cerebral blood flow (CBF) that form the basis of the signals used in functional neuroimaging such as the blood oxygen level-dependent (BOLD) signal. To this end the present study examined the effect of enhanced spike activity per se on CBF in rat cerebellar cortex under conditions of disinhibition, achieved by blocking GABA_A receptors using either bicuculline or picrotoxin. Purkinje cell spiking activity and local field potentials were recorded by glass microelectrodes, and laser Doppler flowmetry was used to monitor CBF. Disinhibition increased Purkinje cell spiking rate to 200–300% of control without incurring any increase in basal CBF. This demonstrates that increased spike activity per se is not sufficient to affect basal CBF. The neurovascular coupling between excitatory synaptic activity and CBF responses evoked by inferior olive (climbing fibre) stimulation was preserved during disinhibition. Thus, the unchanged basal CBF in the presence of the dramatic rise in Purkinje cell spiking rate was not explained by impaired synaptic activity–CBF coupling. On the basis of our previous and the present studies, we conclude that increased spiking activity of principal neurons is neither sufficient nor necessary to elicit CBF responses and in turn BOLD signals, and that activation-dependent vascular signals reflect excitatory synaptic activity.     

Non-uniform olivocerebellar conduction time in the vermis of the rat cerebellum

It has been proposed that the conduction velocities of cerebellar climbing fibre (olivocerebellar) axons are tuned according to length, in order to precisely fix the conduction time between the inferior olive and cerebellar cortex. Some data conflict with this view. We have re-evaluated this issue using the climbing fibre reflex. The white matter of the tip of one folium in lobule VI or VII was stimulated electrically 0.5–1 mm below the surface and recordings were made from Purkinje cells in lobules VIII and IX. Reflex evoked climbing fibre (CF) responses (33 units) were recorded at different depths from Purkinje cells found in a narrow sagittal zone of cortex as complex spikes. The responses had latencies ranging from 4.3 ms to 11.3 ms. A consistent trend was that Purkinje cell responses recorded at greater depth had shorter CF reflex latencies than those recorded more superficially, both in individual experiments and in grouped data. These data show that the CF reflex latency is not constant, but is directly proportional to the distance an action potential has to travel along a CF. These data are not consistent with tuning of CF conduction velocities to normalize olivocerebellar conduction time, but are consistent with a CF conduction velocity in the cortex of approximately 0.6 m s⁻¹. This suggests that climbing fibres projecting to different parts of the cerebellar cortex may have differences in spike conduction time of a few milliseconds, and that submillisecond precision is not an important element of the climbing fibre signal.     

Responses in the inferior olive to stimulation of the cerebellar and cerebral cortices in the cat

1. Extracellular field potentials and single unit responses have been recorded from the inferior olive of the cat following stimulation of the surface of the contralateral paramedian lobule of the cerebellum, and of the ipsilateral cerebral cortex. 2. Cerebellar stimulation results in antidromic invasion of inferior olivary neurones via the climbing fibres. These responses are followed by synaptic discharges which may be generated through climbing fibre recurrent collaterals. 3. Precise histological controls have shown that these responses to stimulation of the paramedian lobule are located in the ventral lamella of the principal olive. 4. Unifocal stimulation of the sensori-motor cortex with surface-anodal pulses evokes synaptically generated discharges of neurones in the central lamella, with a latency of 8-9 msec. The area of cortex yielding responses has been mapped at chosen stimulus intensities and the limitations of the maps have been discussed. 5. It has been shown that the initial excitatory responses obtained from either cortex are followed by an inhibition which lasts about 100 msec, and gives way to a period of recovery or facilitation. This, in turn, is succeeded by a further period of inhibition. Possible neural substrates for these changes have been discussed.     

Metabolic activity of intracerebellar nuclei in the rat: Effects of inferior olive inactivation

Summary Metabolic activity of the intracerebellar nuclei during cryoinactivation of the inferior olive was studied in the anaesthetized rat by using the ¹⁴C-2-deoxyglucose method. Single unit recording of Purkinje cells was simultaneously monitored in the cerebellar cortex.Local inactivation in the inferior olive resulted in regional suppression of complex spike discharges in the cerebellar cortex.An increased metabolic activity was observed in the cerebellar nuclei contralateral to the cryoinactivation site correlating the somatotopically arranged olivo-cerebello-nuclear circuit. This increase was shown to be due specifically to inactivation of the inferior olive, since it was not obtained in a rat in which the inferior olive was previously destroyed by neurotoxic doses of 3-acetylpyridine.The results are interpreted as being due to an increased presynaptic activity of the terminals of the Purkinje cells which fire simple spikes at high rates after climbing fibre deafferentation.     

Actions of acetylcholinesterase in the guinea-pig cerebellar cortex in vitro.

Acetylcholinesterase is released in a calcium-dependent manner when afferents of the cerebellar cortex are stimulated. Since cholinergic transmission is probably insignificant in the cerebellar cortex, the esterase itself might serve as a transmitter or modulator. Therefore, the effect of acetylcholinesterase in the cerebellum was investigated in slices of guinea-pig cerebella during intracellular recording from Purkinje cell somata or dendrites. Addition of acetylcholinesterase (20 U/ml) to the superfusion medium did not change the membrane potential or the input resistance of the Purkinje cells. Thus, esterase does not act like a classical transmitter. The threshold for Na+ spikes generated by intracellular current injection was unaffected, but the threshold for Ca2+ spikes was increased. This increase was abolished by tetrodotoxin (1 microM). Furthermore, when Ca2+ currents were blocked by substituting Mn2+ for Ca2+ (2 mM) a decrease in a Na+ plateau potential was seen in the presence of esterase. The effect of acetylcholinesterase of Ca2+ spikes is therefore most likely due to a reduction of the non-inactivating Na+ current of the Purkinje cell membrane. When present this current contributes to activation of Ca2+ spikes in dendrites. Acetylcholinesterase also enhanced the response of Purkinje cells to the excitatory amino acids glutamate and aspartate thought to be transmitters in the cerebellar cortex. The responses became larger and faster in the presence of esterase. Responses to climbing fibre stimulation were also enhanced by acetylcholinesterase. The late part of this synaptic response was increased. The potentiation by esterase of responses of Purkinje cells to excitatory amino acids and to climbing fibre stimulation may be mediated through interference with transmitter uptake, because it was prevented by treatment with DL-2-amino-4-phosphonobutyric acid (0.5 mM) and di-hydrokainate (0.1 mM). None of the effects of esterase was due to hydrolysis of acetylcholine because irreversible inhibition of the catalytic site of the enzyme with soman did not prevent the actions. The observations were specific for acetylcholinesterase. Butyrylcholinesterase (20-40 U/ml) showed none of the effects. It is concluded that acetylcholinesterase in the cerebellar cortex seems to mediate a novel type of modulation by two separate mechanisms. Esterase reduces the tendency towards Ca2+ spike generation in Purkinje cells. Ca2+ spikes are followed by afterhyperpolarizations and in their absence firing of Na+ spikes at higher frequencies is possible. Secondly, there is an enhancement of the action of excitatory transmitters so that the extended operating range can be utilized.     

Developmental changes in the localisation of the mGluR1alpha subtype of metabotropic glutamate receptors in Purkinje cells.

The regulation of neurotransmitter receptors during synapse formation has been studied extensively at the neuromuscular junction, but little is known about the development of excitatory neurotransmitter receptors during synaptogenesis in central synapses. In this study we show qualitatively and quantitatively that a receptor undergoes changes in localisation on the surface of rat Purkinje cells during development in association with its excitatory synapses. The presence of mGluR1alpha at parallel and climbing fibre synapses on developing Purkinje cells was studied using high-resolution immunoelectron microscopy. Immunoreactivity for mGluR1alpha was detected from embryonic day 18 in Purkinje cells, and showed dramatic changes in its localisation with age. At early postnatal ages (P0 and P3), mGluR1alpha was found both in somata and stem dendrites but was not usually associated with synaptic contacts. At P7, mGluR1alpha became concentrated in somatic spines associated with climbing fibres and in the growing dendritic arborisation even before innervation by parallel fibres. During the second and third postnatal week, when spines and parallel fibre synapses were generated, mGluR1alpha became progressively concentrated in the molecular layer, particularly in the synaptic specialisations. As a result, during the fourth postnatal week, the pattern and level of mGluR1alpha expression became similar to the adult and mGluR1alpha appeared in high density in perisynaptic sites. Our results indicate that mGluR1alpha is present in the developing Purkinje cells prior to their innervation by climbing and parallel fibres and demonstrate that this receptor undergoes a dynamic and specific regulation during postnatal development in association with the establishment of synaptic inputs to Purkinje cell.     

Developmental changes in evoked Purkinje cell complex spike responses

The development of synaptic interconnections between the cerebellum and inferior olive, the sole source of climbing fibers, could contribute to the ontogeny of certain forms of motor learning (e.g., eyeblink conditioning). Purkinje cell complex spikes are produced exclusively by climbing fibers and exhibit short- and long-latency activity in response to somatosensory stimulation. Previous studies have demonstrated that evoked short- and long-latency complex spikes generally occur on separate trials and that this response segregation is regulated by inhibitory feedback to the inferior olive. The present experiment tested the hypothesis that complex spikes evoked by periorbital stimulation are regulated by inhibitory feedback from the cerebellum and that this feedback develops between postnatal days (PND) 17 and 24. Recordings from individual Purkinje cell complex spikes in urethan-anesthetized rats indicated that the segregation of short- and long-latency evoked complex spike activity emerges between PND17 and PND24. In addition, infusion of picrotoxin, a GABAA-receptor antagonist, into the inferior olive abolished the response pattern segregation in PND24 rats, producing evoked complex spike response patterns similar to those characteristic of younger rats. These data support the view that cerebellar feedback to the inferior olive, which is exclusively inhibitory, undergoes substantial changes in the same developmental time window in which certain forms of motor learning emerge.     

Synaptic actions of peripheral nerve impulses upon Deiters neurones via the climbing fibre afferents

1. The cerebellar integration of sensory inputs to Deiters neurones was studied in cats under Nembutal anaesthesia.2. Stimulation of peripheral nerves produced in the Deiters neurones a sequence of an initial excitatory post-synaptic potential (e.p.s.p.) and a later inhibitory post-synaptic potential (i.p.s.p.), or a relatively small e.p.s.p.3. The Deiters neurones were classified as forelimb (FL)- or hind limb (HL)-type cells according to the location of the most effective peripheral nerve. In the FL cells stimulation of the forelimb nerves produced the e.p.s.p.-i.p.s.p. sequence (dominant response), while stimulation of the hind limb nerves was ineffective or produced the small e.p.s.p. (non-dominant response). In contrast, in the HL cells the non-dominant response was evoked from the forelimb nerves, and the dominant response from the hind limb nerves.4. The stimulus intensity-response relation indicates that Group I and II muscle afferents and low and high threshold cutaneous afferents contribute to the dominant and non-dominant responses.5. Antidromic identification of these Deiters neurones revealed that 90% of the HL cells and 85% of the FL cells project to the lumbo-sacral and cervico-thoracic segments of the spinal cord, respectively, while 10% of the HL cells and 15% of the FL cells innervate the cervico-thoracic and lumbo-sacral segments, respectively.6. The mean latency of the e.p.s.p. evoked from the forelimb nerves was 14 msec in the FL cells and 13 msec in the HL cells, and the latency of the e.p.s.p. evoked from the hind limb nerves was 17 msec in the FL cells and 18 msec in the HL cells. The later i.p.s.p. regularly followed the onset of the e.p.s.p. with a delay of 3-5 msec.7. The dominant and non-dominant responses in both types of cells exhibited the following three characteristic features: (i) a strong depression after conditioning stimulation of the inferior olive, (ii) an increase of the inferior olivary excitability during the responses, and (iii) a striking frequency depression with stimulation at relatively low frequency (5-10/sec).8. Consequently it was concluded that all of the responses were produced through the climbing fibres originating from the inferior olive, the i.p.s.p.s due to inhibition from Purkyne cells activated by the climbing fibres and the e.p.s.p.s due to excitation from the collaterals of the climbing fibres.     

The localisation of urocortin in the adult rat cerebellum: a light and electron microscopic study

Light and electron microscopic immunocytochemistry was used to identify the cellular and subcellular localisation of urocortin in the adult rat cerebellum. Urocortin immunoreactivity (UCN-ir) was visualised throughout the cerebellum, yet predominated in the posterior vermal lobules, especially lobules IX and X, the flocculus, paraflocculus and deep cerebellar nuclei. Cortical immunoreactivity was most evident in the Purkinje cell layer and molecular layer. Reaction product, though sparse, was found in the somata of Purkinje cells, primarily in the region of the Golgi apparatus. Purkinje cell dendritic UCN-ir was compartmentalised, with it being prevalent in proximal regions especially where climbing fibres synapsed, yet absent in distal regions where parallel fibres synapsed. In the Purkinje cell layer, the labelling was also contained in axonal terminals, synapsing directly on Purkinje cell somata. These were identified as axon terminals of basket cells based on their morphology. Terminals of stellate cells in the upper molecular layer also expressed the peptide. Whilst somata of inferior olivary neurones showed intense immunoreactivity, axonal labelling was indistinct, with only the terminals of climbing fibres containing reaction product. UCN-ir in the mossy fibre-parallel fibre system was restricted to mossy fibre rosettes of mainly posterior lobules and the varicose terminals of parallel fibres. Furthermore, labelling also was prevalent in glial perikarya and their sheaths.The current study shows, firstly, that urocortin enjoys a close ligand-receptor symmetry in the cerebellum, probably to a greater degree than corticotropin-releasing factor since corticotropin-releasing factor itself is found exclusively in the two major cerebellar afferent systems. Its congregation in excitatory and inhibitory axonal terminals suggests a significant degree of participation in the synaptic milieu, perhaps in the capacity as a neurotransmitter or effecting the release of co-localised neurotransmitters. Finally, its unique distribution in the Purkinje cell dendrite might serve as an anatomical marker of discrete populations of dendritic spines.     

On the Purkinje cell activity increase induced by suppression of inferior olive activity

Summary Previous experiments performed in rats under barbiturate anaesthesia have shown a remarkable increase of simple spike firing rate in cerebellar Purkinje cells following inferior olive lesion or inactivation. The increase is due, at least in part, to the withdrawal of the tonic background activity of corticocerebellar interneurones, which have GABA as a chemical transmitter. Since barbiturates potentiate GABAergic inhibition, it is possible that the effect is due to the presence of this type of anaesthesia. In absence of general anaesthesia, we have compared the simple spike firing rate of the Purkinje cells in rats with intact inferior olive and 3–5 days after inferior olive lesion by 3-acetylpyridine. In the latter condition, the firing rate is significantly higher. In other rats, under urethane anaesthesia, which is not known to interfere with GABAergic transmission, the inferior olive has been reversibly inactivated by applying a cooling probe to the ventral surface of the medulla. Following cooling of the inferior olive on one side, a remarkable increase of simple spike activity, parallel to the disappearance of complex spike activity, has been observed in the Purkinje cells of the contralateral side. These results show that the presence of the simple spike firing increase, which follows the removal of the climbing fibre activity, does not depend on an anaesthetic which potentiates GABAergic transmission, although its amplitude is affected by the same anaesthetic. They suggest, therefore, that the tonic inhibition exerted by the olivocerebellar pathway on the Purkinje cells operates also in physiological conditions. By analysing the pattern of discharge of the Purkinje cell simple spikes in intact rats and following suppression of inferior olive activity, we have seen that, in the latter condition, the highest firing rate is accompanied by a higher degree of regularity. This change of pattern does not depend significantly on the removal of an irregularity induced by the complex spikes, but it is mainly the consequence of the firing rate increase.     

Ultrastructure of synapses in the cerebellar cortex after long-term activation of climbing fibres

Adult male rats were treated for 24 or 48 h with harmaline which selectively activates neurones in the inferior olivary nucleus which give rise to climbing fibres projecting to the cerebellar vermis. Electrophysiological studies have shown that harmaline-induced climbing fibre activity completely blocks the responses of the Purkinje cell to parallel fibre input. Morphometric analysis of the ultrastructure of climbing and parallel fibre synapses revealed no significant differences in morphology between vermis (experimental) and hemisphere (control). These findings indicate that the decreased responsiveness of Purkinje cells to parallel fibre inputs induced by increased climbing fibre activity over 24 or 48 h is not accompanied by any observable structural changes in the cerebellar cortex.     

Early expression of AMPA receptors and lack of NMDA receptors in developing rat climbing fibre synapses

Whether nascent glutamatergic synapses acquire their AMPA receptors constitutively or via a regulated pathway triggered by pre-existing NMDA receptor activation is still an open issue. Here, we provide evidence that some glutamatergic synapses develop without expressing NMDA receptors. Using immunocytochemistry, we showed that synapses between developing rat climbing fibres and Purkinje cells expressed GluR2-containing AMPA receptors as soon as they were formed (i.e. on embryonic day 19) but never carried detectable NMDA receptors. This was confirmed by electrophysiological recordings. Excitatory synaptic currents were recorded in Purkinje cells as early as P0. However, no NMDA receptor-mediated component was found in either spontaneous or evoked synaptic responses. In addition, we ruled out a possible role of extrasynaptic NMDA receptors by showing that AMPA receptor clustering at nascent climbing fibre synapses was not modified by chronic in utero NMDA receptor blockade.     

Effects of afferent volleys from the limbs on the discharge patterns of interpositus neurones in cats anaesthetized with alpha-chloralose.

1. In cats anaesthetized with alpha-chloralose, micro-electrodes have been used to record the discharge patterns of single neurones in the region of the nucleus interpositus. 2. Almost all cells tested could be antidromically invaded following electrical stimulation of the contralateral red nucleus, showing that they were cerebellar efferent neurones. 3. A little over half of the interpositus neurones were spontaneously active, usually at rates of less than 20 impulses/sec. 4. About 40% of the cells had no spontaneous activity, although they gave brisk responses to electrical stimulation of cutaneous nerves. Such silent units were encountered most frequently in the earlier stages of an experiment, but a number were found more than 15 hr after the beginning of an experiment. 5. Stimulation of cutaneous and mixed nerves of the fore and hind limbs provoked impulse discharges of the cells and also produced phases of deceleration of the resting discharge of spontaneously firing cells. 6. The typical response of an interpositus neurone consisted of a short latency (6-35 msec) discharge, usually separated from a long latency (50-500 msec) discharge by a period of inhibition or return to the resting discharge rate. The two phases of excitation appeared to be independently generated, since in a number of cells one phase appeared without the other. In addition, the later phase of excitation was abolished in all cells tested by a small dose of pentobarbitone which produced very little effect on the earlier phase. The long latency response was quantitatively much greater, sometimes consisting of 50 or more impulses in a response which lasted several hundred msec, but was very variable from one trial to another. 7. The long latency discharge and sometimes the preceding inhibition could readily be mimicked by single shock stimulation of the region of the contralateral inferior olive. Short latency discharges were, however, rarely evoked by olivary stimulation. 8. It is suggested that the short latency responses of the interpositus neurones were a result of synaptic excitation via cerebellar afferents, while the ensuing inhibition was a result of post-synaptic inhibition resulting from the Purkinje cell excitation due to the afferent volleys. It is suggested that the long latency excitation is due to the afferent volleys. It is suggested that the long latency excitation is due at least in part to disinhibition resulting from long pauses in Purkinje cell firing following their activation by climbing fibre afferents. 9. The possibility that these long latency responses have a physiological significance in relation to locomotion is discussed.     

Fluctuation of extracellular potassium and calcium in the cerebellar cortex related to climbing fiber activity

It is known that repetitive stimulation of spino- and cortico-olivary pathways can lead to waxing and waning of the evoked climbing fiber responses of cerebellar Purkinje cells. Extracellular recordings performed with ion-selective microelectrodes showed that the amplitude fluctuations of climbing fiber-related field potentials were accompanied by corresponding changes in ion-activity levels. When the amplitude of the field potentials increased, potassium activity rose by up to 0.7 mmol.l^?1 and calcium activity fell by up to 0.3 mmol.l^?1. Ion activities returned to baseline values when climbing fiber responses failed.The rhythmic fluctuations in ion activities have cycle lengths of 10–20 s, or multiples of it. Presumably they are secondary consequences of slow variations of synaptic transmission in the inferior olive. The generator for the slow fluctuations of olivary synaptic transmission and the consequent variations in ion activities within the cerebellum is probably located within the inferior olive. The ionic modulations could have functional implications for cerebellar information processing. Possible mechanisms of the rhythmogenesis are discussed.     

A theory of cerebellar cortex

1. A detailed theory of cerebellar cortex is proposed whose consequence is that the cerebellum learns to perform motor skills. Two forms of input-output relation are described, both consistent with the cortical theory. One is suitable for learning movements (actions), and the other for learning to maintain posture and balance (maintenance reflexes).2. It is known that the cells of the inferior olive and the cerebellar Purkinje cells have a special one-to-one relationship induced by the climbing fibre input. For learning actions, it is assumed that:(a) each olivary cell responds to a cerebral instruction for an elemental movement. Any action has a defining representation in terms of elemental movements, and this representation has a neural expression as a sequence of firing patterns in the inferior olive; and(b) in the correct state of the nervous system, a Purkinje cell can initiate the elemental movement to which its corresponding olivary cell responds.3. Whenever an olivary cell fires, it sends an impulse (via the climbing fibre input) to its corresponding Purkinje cell. This Purkinje cell is also exposed (via the mossy fibre input) to information about the context in which its olivary cell fired; and it is shown how, during rehearsal of an action, each Purkinje cell can learn to recognize such contexts. Later, when the action has been learnt, occurrence of the context alone is enough to fire the Purkinje cell, which then causes the next elemental movement. The action thus progresses as it did during rehearsal.4. It is shown that an interpretation of cerebellar cortex as a structure which allows each Purkinje cell to learn a number of contexts is consistent both with the distributions of the various types of cell, and with their known excitatory or inhibitory natures. It is demonstrated that the mossy fibre-granule cell arrangement provides the required pattern discrimination capability.5. The following predictions are made.(a) The synapses from parallel fibres to Purkinje cells are facilitated by the conjunction of presynaptic and climbing fibre (or post-synaptic) activity.(b) No other cerebellar synapses are modifiable.(c) Golgi cells are driven by the greater of the inputs from their upper and lower dendritic fields.6. For learning maintenance reflexes, 2(a) and 2(b) are replaced by2'. Each olivary cell is stimulated by one or more receptors, all of whose activities are usually reduced by the results of stimulating the corresponding Purkinje cell.7. It is shown that if (2') is satisfied, the circuit receptor --> olivary cell --> Purkinje cell --> effector may be regarded as a stabilizing reflex circuit which is activated by learned mossy fibre inputs. This type of reflex has been called a learned conditional reflex, and it is shown how such reflexes can solve problems of maintaining posture and balance.8. 5(a), and either (2) or (2') are essential to the theory: 5(b) and 5(c) are not absolutely essential, and parts of the theory could survive the disproof of either.