Characteristic patterns of EEG oscillations in sheep (Ovis aries) induced by ketamine may explain the psychotropic effects seen in humans.

Ketamine is a valuable anaesthetic and analgesic that in recent years has gained notoriety as a recreational drug. Recently, ketamine has also been proposed as a novel treatment for depression and post-traumatic stress disorder. Beyond its anaesthetic actions, however, the effects of ketamine on brain activity have rarely been probed. Here we examined the cortical electroencephalography (EEG) response to ketamine of 12 sheep. Following ketamine administration, EEG changes were immediate and widespread, affecting the full extent of the EEG frequency spectrum measured (0-125 Hz). After recovery from sedation during which low frequency activity dominated, the EEG was characterised by short periods (2-3 s) of alternating low (<14 Hz) and high (>35 Hz) frequency oscillation. This alternating EEG rhythm phase is likely to underlie the dissociative actions of ketamine, since it is during this phase that ketamine users report hallucinations. At the highest intravenous dose used (24 mg/kg), in 5/6 sheep we observed a novel effect of ketamine, namely the complete cessation of cortical EEG activity. This persisted for up to several minutes, after which cortical activity resumed. This phenomenon is likely to explain the 'k-hole', a state of oblivion likened to a near death experience that is keenly sought by ketamine abusers.

Techniques for chronic monitoring of brain activity in freely moving sheep using wireless EEG recording.

BACKGROUND: Large mammals with complex central nervous systems offer new possibilities for translational research into basic brain function. Techniques for monitoring brain activity in large mammals, however, are not as well developed as they are in rodents. NEW METHOD: We have developed a method for chronic monitoring of electroencephalographic (EEG) activity in unrestrained sheep. We describe the methods for behavioural training prior to implantation, surgical procedures for implantation, a protocol for reliable anaesthesia and recovery, methods for EEG data collection, as well as data pertaining to suitability and longevity of different types of electrodes. RESULTS: Sheep tolerated all procedures well, and surgical complications were minimal. Electrode types used included epidural and subdural screws, intracortical needles and subdural disk electrodes, with the latter producing the best and most reliable results. The implants yielded longitudinal EEG data of consistent quality for periods of at least a year, and in some cases up to 2 years. COMPARISON WITH EXISTING METHODS: This is the first detailed methodology to be described for chronic brain function monitoring in freely moving unrestrained sheep. CONCLUSIONS: The developed method will be particularly useful in chronic investigations of brain activity during normal behaviour that can include sleep, learning and memory. As well, within the context of disease, the method can be used to monitor brain pathology or the progress of therapeutic trials in transgenic or natural disease models in sheep.

Spike sorting based upon machine learning algorithms (SOMA).

We have developed a spike sorting method, using a combination of various machine learning algorithms, to analyse electrophysiological data and automatically determine the number of sampled neurons from an individual electrode, and discriminate their activities. We discuss extensions to a standard unsupervised learning algorithm (Kohonen), as using a simple application of this technique would only identify a known number of clusters. Our extra techniques automatically identify the number of clusters within the dataset, and their sizes, thereby reducing the chance of misclassification. We also discuss a new pre-processing technique, which transforms the data into a higher dimensional feature space revealing separable clusters. Using principal component analysis (PCA) alone may not achieve this. Our new approach appends the features acquired using PCA with features describing the geometric shapes that constitute a spike waveform. To validate our new spike sorting approach, we have applied it to multi-electrode array datasets acquired from the rat olfactory bulb, and from the sheep infero-temporal cortex, and using simulated data. The SOMA sofware is available at http://www.sussex.ac.uk/Users/pmh20/spikes.

Applications of multi-variate analysis of variance (MANOVA) to multi-electrode array electrophysiology data.

We have developed an adaptation of multi-variate analysis of variance (MANOVA) to analyze statistically both local and global patterns of multi-electrode array (MEA) electrophysiology data where the activities of many (typically >100) neurons have been recorded simultaneously. Whereas simple application of standard MANOVA techniques prohibits extraction of useful information in this kind of data, our new approach, MEANOVA (=MEA+MANOVA), allows a more useful and powerful approach to analyze such complex neurophysiological data. The MEANOVA test enables the detection of the "hot-spots" in the MEA data and has been validated using recordings from the rat olfactory bulb. To further validate the power of this approach, we have also applied the MEANOVA test to data obtained from a simple computational network model. This MEANOVA software and other useful statistical methods for MEA data can be downloaded from http://www.sussex.ac.uk/Users/pmh20

Tracking memory's trace.

There is strong converging evidence that the intermediate and medial part of the hyperstriatum ventrale of the chick brain is a memory store for information acquired through the learning process of imprinting. Neurons in this memory system come, through imprinting, to respond selectively to the imprinting stimulus (IS) neurons and so possess the properties of a memory trace. Therefore, the responses of the intermediate and medial part of the hyperstriatum ventrale neurons to a visual imprinting stimulus were determined before, during, and after training. Of the total recorded population, the proportions of IS neurons shortly after each of two 1-h training sessions were significantly higher (approximately 2 times) than the pretraining proportion. However, approximately 4.5 h later this proportion had fallen significantly and did not differ significantly from the pretraining proportion. Nevertheless, approximately 21.5 h after the end of training, the proportion of IS neurons was at its highest (approximately 3 times the pretraining level). No significant fluctuations occurred in the proportions of neurons responding to the alternative stimulus. In addition, nonmonotonic changes were found commonly in the activity of 230 of the neurons tracked individually from before training to shortly after the end of training. Thus the pattern of change in responsiveness both at the population level and at the level of individual neurons was highly nonmonotonic. Such a pattern of change is not consistent with simple models of memory based on synaptic strengthening to asymptote. A model is proposed that accounts for the changes in the population responses to the imprinting stimulus in terms of changes in the responses of individual neurons.

Is neuronal encoding of subject-object distance dependent on learning?

Recordings were made in the intermediate and medial part of the hyperstriatum ventrale of behaving domestic chicks which had been imprinted (trained) by being exposed to a training stimulus. Neurons were tested for responsiveness to the training stimulus and to an alternative stimulus at each of three distances (d = 0.5 m, 1 m, 2 m) from the chick. For responses to the training stimulus 24/78 (31%) responsive neurons were d-sensitive, i.e. responses changed with distance. For responses to the alternative stimulus, a similar proportion of neurons was d-sensitive (16/57, 28%). Six d-invariant neurons responded similarly at each distance: four to the training and two to the alternative stimulus. Thus no effect of learning on d-sensitive or d-invariant neuronal responsiveness was found.

The recognition memory of imprinting: biochemistry and electrophysiology.

A restricted part of the intermediate and medial part of the hyperstriatum ventrale (IMHV) of the domestic chick forebrain is pivotal to the learning process of imprinting and is probably the site at which information about an imprinting stimulus is stored. A range of learning-related changes occur in the IMHV between 1 and 24 h after training. The earliest change described is in Fos-like immunoreactivity. There follow changes in phosphorylation of the protein kinase C substrate MARCKS, morphological changes in axospinous synapses, an increase in NMDA receptor number and increases in amounts of the major isoforms of the neural cell adhesion molecule and clathrin heavy chain. All but the change in Fos-immunopositivity occurs in the left, but not the right, IMHV. Insufficient nitric oxide synthase is available in the IMHV to support the hypothesis that nitric oxide is a retrograde messenger contributing to the effect on Fos-like immunoreactivity. In chicks anaesthetised approximately 24 h after imprinting training, the spontaneous mean neuronal firing rate is related to a preference score (a measure of learning). In unanaesthetised chicks 24 h after training, the responsiveness of some IMHV neurons is biassed specifically towards the imprinting stimulus.The responses of other neurons in the IMHV generalise across some features of the training stimulus, such as form or colour. Some neurons in the IMHV of unanaesthetised chicks are responsive to the distance of an imprinting stimulus from the chick; distance-sensitive neurons can be distinguished from distance-insensitive neurones by the action potential shape.

Short communication: hippocampal neuronal activity and imprinting in the behaving domestic chick.

The hippocampus of the chick projects to the intermediate and medial part of the hyperstriatum ventrale (IMHV) which stores information acquired through the learning process of imprinting. We have investigated whether the response properties of hippocampal neurons are similar to those of IMHV neurons. Chicks were imprinted by exposure, one group (n = 7) to a rotating red box (RB), the other (n = 5) to a rotating blue cylinder (BC). Four chicks were untrained. The following day, when the chicks were approximately 48 h old, neuronal activity was recorded in the left hippocampus. The proportion of neurons responding to the RB and that to the BC in untrained chicks were compared with the proportions in trained birds. (i) In RB-trained chicks both the proportion responding to the RB and that to the BC were significantly increased. (ii) In BC-trained chicks no significant effect on these proportions was found. Of the responsive neurons some were colour (red or blue) sensitive and others were shape (box or cylinder) sensitive; the proportions so responsive were not influenced by training condition. Certain neurons responded significantly differently when a stimulus was 0.5 m or 2 m from the chick (35%; d-sensitive); very few neurons were equivalently responsive to a stimulus at both distances (3%; d-invariant). These proportions were not significantly affected by training condition. Hippocampal responses are compared with those in the left IMHV. It is concluded that IMHV responses do not passively reflect those of hippocampal neurons.

Neural encoding of subject-object distance in a visual recognition system.

Domestic chicks follow a familiar (imprinted) object when it recedes. In behaving, imprinted chicks with no experience of objects at different distances, neuronal activity was recorded from the intermediate and medial part of the hyperstriatum ventrale (IMHV), a brain region crucial for the recognition memory underlying imprinting. We found that (i) some neurones responded equivalently, irrespective of the subject-object distance d (d-invariant); (ii) other neurones responded differently at different values of d (d-sensitive); (iii) these response types were found in monocular chicks and in chicks with both eyes exposed; (iv) the action potential shape of d-invariant neurones was different from that of other neurones and (v) the spontaneous firing rate of some neurones was correlated with locomotor activity. Taken together with previous findings, the results raise the possibility that IMHV has a major role to play in the sensory and motor-control aspects of imprinting in addition to its mnemonic functions.

Neurophysiological investigations of a recognition memory system for imprinting in the domestic chick.

The responsiveness of neurons in a region of the chick brain involved in the learning process of imprinting, the right intermediate and medial hyperstriatum ventrale (right IMHV), has been investigated in unanaesthetized, trained and untrained chicks. The results demonstrate that neuronal responsiveness in this region reflects a variety of behavioural consequences of imprinting and is markedly altered as a result of the learning process. Groups of chicks (nine in each group) were either dark-reared or trained (imprinted) by exposure to a rotating red box or a rotating blue cylinder. Recordings of single or small groups of neurons were subsequently made from 156 sites in the right IMHV while the 2-day-old chicks were free to move in a running wheel. There was a highly significant increase in the proportion of sites responsive to the stimulus used to train the birds compared to the proportion responsive to that stimulus in dark-reared birds (30 and 9% respectively). These changes were found when either the red box or the blue cylinder was used to train the bird, the changes being similar for both stimuli. There was also a significant increase in the mean magnitude of the change in neuronal activity on stimulus presentation for the training stimulus compared to the same stimulus when not used in the bird's training. No significant effects of the training experience of the chicks were found upon either the magnitude of evoked activity or the proportion of sites responsive to a rotating stuffed jungle fowl or the sound of the maternal call. The presence of the training stimulus was selectively signalled by the response at certain sites. At other sites there was response generalization across stimulus shape or colour. A comparison with results for the left IMHV demonstrates both similarities and differences in neuronal responsiveness between the two regions. In both regions imprinting selectively enhances neuronal responsiveness to the training stimulus. However, for trained birds the mean proportion of sites responding to whichever of the red box or the blue cylinder was not used in the bird's training was significantly lower in the right than the left IMHV. These results are discussed in relationship to previously reported asymmetries in the response of the right and left IMHV regions to imprinting. A model is introduced to explain the physiological findings. The effects of training on right IMHV neuronal function are consistent with a long-term role for this region in the recognition memory of imprinting.

Sustained potential shifts in the toad tectum reflect prey-catching and avoidance behavior.

Sustained potential shifts (SPS) were recorded for 10 s from the surface of the optic tectum of toads presented with live prey and moving artificial prey stimuli. On the anterior tectal surface, a negative SPS was followed by a positive wave; the converse was true for the posterior tectum. Some animals were immobilized, and they exhibited a monophasic negative SPS in the anterior tectum and a positive wave in more posterior regions. The number of orienting responses made by toads to moving "wormlike" stimuli was reflected in the amplitude of the SPS, as was avoidance to stimuli in an "antiworm" configuration. Behavioral activity was most closely related to the negativity of the SPS recording. The SPS of toads responding to live prey showed no direct time relationship between the SPS and behavior, suggesting that the SPS reflects sensory or decision-making activity rather than the consequent behavior.

Sustained potential shift responses and their relationship to the ECG response during arousal in the goldfish (Carassius auratus).

1. Goldfish, when presented with a 10 sec light-on stimulus against a background of 2 hr of sensory restriction, exhibited sustained potential shift (SPS) activity, of differing polarity, at each of four surface recording sites, on the medulla, cerebellum, optic tectum and telencephalon. 2. Principle components analysis (PCA) indicated that SPS responses from each region comprised superimposed early and late waveforms. At the cerebellar, tectal and telencephalic surfaces, neuronal activity appeared to contribute strongly to the early (less than 2 sec) SPS waveform. 3. While, in response to repeated stimulus presentations, habituation was apparent in the early SPS waveforms recorded from the medulla, cerebellum and telencephalon, an increase in negativity occurred in late SPS waveforms throughout the brain. 4. The tectal SPS response was directly proportional to the telencephalic SPS response both in terms of average SPS amplitudes following the first presentation of the light-on stimulus and in terms of their increasing negativity in response to stimulus repetition. 5. The increasing negativity of the telencephalic SPS was also associated with the habituation of the ECG response over repeated trials. 6. Results are discussed with regard to a possible neuromodulatory role for glia.

Dendritic and sustained shifts in potential to electrical stimulation of the anuran tectal surface.

1. Recordings of dendritic potentials and sustained potential shifts (SPS) were made from the brain of immobilised frogs during surface tectal electrical stimulation. 2. Single pulses evoked dendritic responses; trains caused decay of dendritic responses on the background of the evoked SPS. 3. The tectal surface SPS declined with distance from the stimulating electrode. 4. The negative surface SPS declined with tectal depth to ca 300 microns, then reversed polarity and increased in amplitude with depth up to 700 microns.