Neuronal dynamics supporting formation and recombination of cross-modal olfactory-tactile association in the rat hippocampal formation.

The present study is aimed at describing some aspects of the neural dynamics supporting discrimination of olfactory-tactile paired-associated stimuli during acquisition of new pairs and during recombination of previously learned pairs in the rat. To solve the task, animals have to identify one odor-texture (OT) combination associated with a food reward among three cups with overlapping elements. Previous experiments demonstrated that the lateral entorhinal cortex (LEC) is involved in the processes underlying OT acquisition, whereas the dorsal hippocampus (DH) is selectively involved in the recombination processes. In the present study, local field potentials were recorded form the anterior piriform cortex (aPC), LEC, and DH in freely moving rats performing these tasks. Signal analysis focused on theta (5-12 Hz)- and beta-band (15-40 Hz) oscillatory activities in terms of both amplitude and synchrony. The results show that cue sampling was associated with a significant increase in the beta-band activity during the choice period in both the aPC and the LEC, and is modulated by level of expertise and the animal's decision. In addition, this increase was significantly higher during the recombination compared with the acquisition of the OT task, specifically when animals had to neglect the odor previously associated with the reward. Finally, a significant decrease in coherence in the theta band between LEC and DH was observed in the recombination but not in the acquisition task. These data point to specific neural signatures of simple and complex cross-modal sensory processing in the LEC-DH complex. NEW & NOTEWORTHY This study is the first to describe electrophysiological correlates of cross-modal olfactory-tactile integration in rats. Recordings were sought from the lateral entorhinal cortex and the dorsal hippocampus because previous studies have shown their role in the formation and in the recombination of previously learned associations. We identified specific oscillatory-evoked neural responses in these structures in the theta and beta bands, which characterize acquisition and recombination of cross-modal olfactory-tactile pairs.

Brainstem mechanisms of paradoxical (REM) sleep generation.

Paradoxical sleep (PS) is characterized by EEG activation with a disappearance of muscle tone and the occurrence of rapid eye movements (REM) in contrast to slow-wave sleep (SWS, also known as non-REM sleep) identified by the presence of delta waves. Soon after the discovery of PS, it was demonstrated that the structures necessary and sufficient for its genesis are restricted to the brainstem. We review here recent results indicating that brainstem glutamatergic and GABAergic, rather than cholinergic and monoaminergic, neurons play a key role in the genesis of PS. We hypothesize that the entrance to PS from SWS is due to the activation of PS-on glutamatergic neurons localized in the pontine sublaterodorsal tegmental nucleus. The activation of these neurons would be due to a permanent glutamatergic input arising from the lateral and ventrolateral periaqueductal gray (vlPAG) and the removal at the onset of PS of a GABAergic inhibition present during W and SWS. Such inhibition would be coming from PS-off GABAergic neurons localized in the vlPAG and the adjacent deep mesencephalic reticular nucleus. The cessation of activity of these PS-off GABAergic neurons at the onset and during PS would be due to direct projections from intermingled GABAergic PS-on neurons. Activation of PS would depend on the reciprocal interactions between the GABAergic PS-on and PS-off neurons, intrinsic cellular and molecular events, and integration of multiple physiological parameters.

Improving Stereotaxic Neurosurgery Techniques and Procedures Greatly Reduces the Number of Rats Used per Experimental Group-A Practice Report.

Techniques of stereotaxic surgery are commonly used in research laboratories by a range of students, technicians, and researchers. To meet the evolving requirements imposed by international legislation, and to promote the implementation of 3R rules (replacement, reduction, and refinement) by reducing experimental error, animal morbidity, and mortality, it is essential that standard operating procedures and proper conduct following such complex surgeries be precisely described and respected. The present report shows how refinements of our own neurosurgical techniques over decades, have significantly reduced the number of animals (rats) used in experiments and improved the animals' well-being during the post-surgical recovery period. The current pre-, per-, and post-surgical procedures used in our laboratory are detailed. We describe the practical aspects of stereotaxic neurosurgery that have been refined in our laboratory since 1992 and that cover various areas including appropriate anesthesia and pain management during and after surgery, methods to determine the stereotaxic coordinates, and the best approach to the target brain structure. The application of these optimal surgical methods that combine reliable and reproducible results with an acute awareness of ethics and animal welfare leads to a significant reduction in the number of animals included in experimental research in accordance with ethical and regulatory rules as required by the European Directive on laboratory animal welfare.

Neuronal activity in the primary somatosensory thalamocortical loop is modulated by reward contingency during tactile discrimination.

Delayed-response sensory discrimination is believed to require primary sensory thalamus and cortex for early stimulus identification and higher-order forebrain regions for the late association of stimuli with rewarded motor responses. Here we investigate neuronal responses in the rat primary somatosensory cortex (S1) and ventral posterior medial nucleus of the thalamus (VPM) during a tactile discrimination task that requires animals to associate two different tactile stimuli with two corresponding choices of spatial trajectory to be rewarded. To manipulate reward expectation, neuronal activity observed under regular reward contingency (CR) was compared with neuronal activity recorded during freely rewarded (FR) trials, in which animals obtained reward regardless of their choice of spatial trajectory. Across-trial firing rates of S1 and VPM neurons varied according to the reward contingency of the task. Analysis of neuronal ensemble activity by an artificial neural network showed that stimulus-related information in S1 and VPM increased from stimulus sampling to reward delivery in CR trials but decreased to chance levels when animals performed FR trials, when stimulus discrimination was irrelevant for task execution. Neuronal ensemble activity in VPM was only correlated with task performance during stimulus presentation. In contrast, S1 neuronal activity was highly correlated with task performance long after stimulus removal, a relationship that peaked during the 300 ms that preceded reward delivery. Together, our results indicate that neuronal activity in the primary somatosensory thalamocortical loop is strongly modulated by reward contingency.

Sleep architecture of the melanin-concentrating hormone receptor 1-knockout mice.

Growing amounts of data indicate involvement of the posterior hypothalamus in the regulation of sleep, especially paradoxical sleep (PS). Accordingly, we previously showed that the melanin-concentrating hormone (MCH)-producing neurons of the rat hypothalamus are selectively activated during a PS rebound. In addition, intracerebroventricular infusion of MCH increases total sleep duration, suggesting a new role for MCH in sleep regulation. To determine whether activation of the MCH system promotes sleep, we studied spontaneous sleep and its homeostatic regulation in mice with deletion of the MCH-receptor 1 gene (MCH-R1-/- vs. MCH-R1+/+) and their behavioural response to modafinil, a powerful antinarcoleptic drug. Here, we show that the lack of functional MCH-R1 results in a hypersomniac-like phenotype, both in basal conditions and after total sleep deprivation, compared to wild-type mice. Further, we found that modafinil was less potent at inducing wakefulness in MCH-R1-/- than in MCH-R1+/+ mice. We report for the first time that animals with genetically inactivated MCH signaling exhibit altered vigilance state architecture and sleep homeostasis. This study also suggests that the MCH system may modulate central pathways involved in the wake-promoting effect of modafinil.

Adaptive quantization of local field potentials for wireless implants in freely moving animals: an open-source neural recording device.

OBJECTIVE: Modern neuroscience research requires electrophysiological recording of local field potentials (LFPs) in moving animals. Wireless transmission has the advantage of removing the wires between the animal and the recording equipment but is hampered by the large number of data to be sent at a relatively high rate. APPROACH: To reduce transmission bandwidth, we propose an encoder/decoder scheme based on adaptive non-uniform quantization. Our algorithm uses the current transmitted codeword to adapt the quantization intervals to changing statistics in LFP signals. It is thus backward adaptive and does not require the sending of side information. The computational complexity is low and similar at the encoder and decoder sides. These features allow for real-time signal recovery and facilitate hardware implementation with low-cost commercial microcontrollers. MAIN RESULTS: As proof-of-concept, we developed an open-source neural recording device called NeRD. The NeRD prototype digitally transmits eight channels encoded at 10 kHz with 2 bits per sample. It occupies a volume of 2 × 2 × 2 cm3 and weighs 8 g with a small battery allowing for 2 h 40 min of autonomy. The power dissipation is 59.4 mW for a communication range of 8 m and transmission losses below 0.1%. The small weight and low power consumption offer the possibility of mounting the entire device on the head of a rodent without resorting to a separate head-stage and battery backpack. The NeRD prototype is validated in recording LFPs in freely moving rats at 2 bits per sample while maintaining an acceptable signal-to-noise ratio (>30 dB) over a range of noisy channels. SIGNIFICANCE: Adaptive quantization in neural implants allows for lower transmission bandwidths while retaining high signal fidelity and preserving fundamental frequencies in LFPs.

Long-lasting novelty-induced neuronal reverberation during slow-wave sleep in multiple forebrain areas.

The discovery of experience-dependent brain reactivation during both slow-wave (SW) and rapid eye-movement (REM) sleep led to the notion that the consolidation of recently acquired memory traces requires neural replay during sleep. To date, however, several observations continue to undermine this hypothesis. To address some of these objections, we investigated the effects of a transient novel experience on the long-term evolution of ongoing neuronal activity in the rat forebrain. We observed that spatiotemporal patterns of neuronal ensemble activity originally produced by the tactile exploration of novel objects recurred for up to 48 h in the cerebral cortex, hippocampus, putamen, and thalamus. This novelty-induced recurrence was characterized by low but significant correlations values. Nearly identical results were found for neuronal activity sampled when animals were moving between objects without touching them. In contrast, negligible recurrence was observed for neuronal patterns obtained when animals explored a familiar environment. While the reverberation of past patterns of neuronal activity was strongest during SW sleep, waking was correlated with a decrease of neuronal reverberation. REM sleep showed more variable results across animals. In contrast with data from hippocampal place cells, we found no evidence of time compression or expansion of neuronal reverberation in any of the sampled forebrain areas. Our results indicate that persistent experience-dependent neuronal reverberation is a general property of multiple forebrain structures. It does not consist of an exact replay of previous activity, but instead it defines a mild and consistent bias towards salient neural ensemble firing patterns. These results are compatible with a slow and progressive process of memory consolidation, reflecting novelty-related neuronal ensemble relationships that seem to be context- rather than stimulus-specific. Based on our current and previous results, we propose that the two major phases of sleep play distinct and complementary roles in memory consolidation: pretranscriptional recall during SW sleep and transcriptional storage during REM sleep.

Localization of the neurons active during paradoxical (REM) sleep and projecting to the locus coeruleus noradrenergic neurons in the rat.

Locus coeruleus (LC) noradrenergic neurons are active during wakefulness, slow their discharge rate during slow wave sleep, and stop firing during paradoxical sleep (PS). A large body of data indicates that their inactivation during PS is due to a tonic GABAergic inhibition. To localize the neurons responsible for such inhibition, we first examined the distribution of retrogradely and Fos double-immunostained neurons following cholera toxin b subunit (CTb) injection in the LC of control rats, rats selectively deprived of PS for 3 days, and rats allowed to recover for 3 hours from such deprivation. We found a significant number of CTb/Fos double-labeled cells only in the recovery group. The largest number of CTb/Fos double-labeled cells was found in the dorsal paragigantocellular reticular nucleus (DPGi). It indeed contained 19% of the CTb/Fos double-labeled neurons, whereas the ventrolateral periaqueductal gray (vlPAG) contained 18.3% of these neurons, the lateral paragigantocellular reticular nucleus (LPGi) 15%, the lateral hypothalamic area 9%, the lateral PAG 6.7%, and the rostral PAG 6%. In addition, CTb/Fos double-labeled cells constituted 43% of all the singly CTb-labeled cells counted in the DPGi compared with 29% for the LPGi, 18% for the rostral PAG, and 10% or less for the other structures. Although all these populations of CTb/Fos double-labeled neurons could be GABAergic and tonically inhibit LC neurons during PS, our results indicate that neurons from the DPGi constitute the best candidate for this role.

Paradoxical (REM) sleep genesis: the switch from an aminergic-cholinergic to a GABAergic-glutamatergic hypothesis.

In the middle of the last century, Michel Jouvet discovered paradoxical sleep (PS), a sleep phase paradoxically characterized by cortical activation and rapid eye movements and a muscle atonia. Soon after, he showed that it was still present in "pontine cats" in which all structures rostral to the brainstem have been removed. Later on, it was demonstrated that the pontine peri-locus coeruleus alpha (peri-LCalpha in cats, corresponding to the sublaterodorsal nucleus, SLD, in rats) is responsible for PS onset. It was then proposed that the onset and maintenance of PS is due to a reciprocal inhibitory interaction between neurons presumably cholinergic specifically active during PS localized in this region and monoaminergic neurons. In the last decade, we have tested this hypothesis with our model of head-restrained rats and functional neuroanatomical studies. Our results confirmed that the SLD in rats contains the neurons responsible for the onset and maintenance of PS. They further indicate that (1) these neurons are non-cholinergic possibly glutamatergic neurons, (2) they directly project to the glycinergic premotoneurons localized in the medullary ventral gigantocellular reticular nucleus (GiV), (3) the main neurotransmitter responsible for their inhibition during waking (W) and slow wave sleep (SWS) is GABA rather than monoamines, (4) they are constantly and tonically excited by glutamate and (5) the GABAergic neurons responsible for their tonic inhibition during W and SWS are localized in the deep mesencephalic reticular nucleus (DPMe). We also showed that the tonic inhibition of locus coeruleus (LC) noradrenergic and dorsal raphe (DRN) serotonergic neurons during sleep is due to a tonic GABAergic inhibition by neurons localized in the dorsal paragigantocellular reticular nucleus (DPGi) and the ventrolateral periaqueductal gray (vlPAG). We propose that these GABAergic neurons also inhibit the GABAergic neurons of the DPMe at the onset and during PS and are therefore responsible for the onset and maintenance of PS.

The switch of subthalamic neurons from an irregular to a bursting pattern does not solely depend on their GABAergic inputs in the anesthetic-free rat.

The subthalamic nucleus (STN) powerfully controls basal ganglia outputs and has been implicated in movement disorders observed in Parkinson's disease because of its pathological mixed burst firing mode and hyperactivity. A recent study suggested that reciprocally connected glutamatergic STN and GABAergic globus pallidus (GP) neurons act in vitro as a generator of bursting activity in basal ganglia. In vivo, we reported that GP neurons increased their firing rate in wakefulness (W) compared with slow-wave sleep (SWS) without any change in their random pattern. In contrast, STN neurons exhibited similar firing rates in W and SWS, with an irregular pattern in W and a bursty one in SWS. Thus, the pallidal GABAergic tone might control the STN pattern. This hypothesis was tested by mimicking such variations with microiontophoresis of GABA receptor ligands. GABA agonists specifically decreased the STN firing rate but did not affect its firing pattern. GABA(A) (but not GABA(B)) antagonists strongly enhanced the STN mean discharge rate during all vigilance states up to three to five times its basal activity. However, such applications did not change the typical W random pattern. When applied during SWS, GABA(A) antagonists strongly reinforced the spontaneous bursty pattern into a particularly marked one with instantaneous frequencies reaching 500-600 Hz. SWS-W transitions occurring during ongoing antagonist iontophoresis invariably disrupted the bursty pattern into a random one. Thus GABA(A) receptors play a critical, but not exclusive, role in regulating the excitatory STN influence on basal ganglia outputs.

Global forebrain dynamics predict rat behavioral states and their transitions.

The wake-sleep cycle, a spontaneous succession of global brain states that correspond to major overt behaviors, occurs in all higher vertebrates. The transitions between these states, at once rapid and drastic, remain poorly understood. Here, intracranial local field potentials (LFPs) recorded in the cortex, hippocampus, striatum, and thalamus were used to characterize the neurophysiological correlates of the rat wake-sleep cycle. By way of a new method for the objective classification and quantitative investigation of all major brain states, we demonstrate that global brain state transitions occur simultaneously across multiple forebrain areas as specific spectral trajectories with characteristic path, duration, and coherence bandwidth. During state transitions, striking changes in neural synchronization are effected by the prominent narrow-band LFP oscillations that mark state boundaries. Our results demonstrate that distant forebrain areas tightly coordinate the processing of neural information during and between global brain states, indicating a very high degree of functional integration across the entire wake-sleep cycle. We propose that transient oscillatory synchronization of synaptic inputs, which underlie the rapid switching of global brain states, may facilitate the exchange of information within and across brain areas at the boundaries of very distinct neural processing regimens.

GABAergic control of hypothalamic melanin-concentrating hormone-containing neurons across the sleep-waking cycle.

The perifornical-lateral hypothalamic area is implicated in regulating waking and paradoxical sleep. The blockade of GABAA receptors by iontophoretic applications of bicuculline (or gabazine) into the perifornical-lateral hypothalamic area induced a continuous quiet waking state associated to a robust muscle tone in head-restrained rats. During the effects, sleep was totally suppressed. In rats killed at the end of a 90 min ejection of bicuculline, Fos expression was induced in approximately 28% of the neurons immunoreactive for hypocretin and in approximately 3% of the neurons immunostained for melanin-concentrating hormone within the ejection site. These results suggest that neurons containing melanin-concentrating hormone are not active during waking and that the lack of a potent GABAergic influence during waking is consistent with their role in sleep regulation.

Unsupervised online classifier in sleep scoring for sleep deprivation studies.

STUDY OBJECTIVE: This study was designed to evaluate an unsupervised adaptive algorithm for real-time detection of sleep and wake states in rodents. DESIGN: We designed a Bayesian classifier that automatically extracts electroencephalogram (EEG) and electromyogram (EMG) features and categorizes non-overlapping 5-s epochs into one of the three major sleep and wake states without any human supervision. This sleep-scoring algorithm is coupled online with a new device to perform selective paradoxical sleep deprivation (PSD). SETTINGS: Controlled laboratory settings for chronic polygraphic sleep recordings and selective PSD. PARTICIPANTS: Ten adult Sprague-Dawley rats instrumented for chronic polysomnographic recordings. MEASUREMENTS: The performance of the algorithm is evaluated by comparison with the score obtained by a human expert reader. Online detection of PS is then validated with a PSD protocol with duration of 72 hours. RESULTS: Our algorithm gave a high concordance with human scoring with an average κ coefficient > 70%. Notably, the specificity to detect PS reached 92%. Selective PSD using real-time detection of PS strongly reduced PS amounts, leaving only brief PS bouts necessary for the detection of PS in EEG and EMG signals (4.7 ± 0.7% over 72 h, versus 8.9 ± 0.5% in baseline), and was followed by a significant PS rebound (23.3 ± 3.3% over 150 minutes). CONCLUSIONS: Our fully unsupervised data-driven algorithm overcomes some limitations of the other automated methods such as the selection of representative descriptors or threshold settings. When used online and coupled with our sleep deprivation device, it represents a better option for selective PSD than other methods like the tedious gentle handling or the platform method.

Role of the lateral paragigantocellular nucleus in the network of paradoxical (REM) sleep: an electrophysiological and anatomical study in the rat.

The lateral paragigantocellular nucleus (LPGi) is located in the ventrolateral medulla and is known as a sympathoexcitatory area involved in the control of blood pressure. In recent experiments, we showed that the LPGi contains a large number of neurons activated during PS hypersomnia following a selective deprivation. Among these neurons, more than two-thirds are GABAergic and more than one fourth send efferent fibers to the wake-active locus coeruleus nucleus. To get more insight into the role of the LPGi in PS regulation, we combined an electrophysiological and anatomical approach in the rat, using extracellular recordings in the head-restrained model and injections of tracers followed by the immunohistochemical detection of Fos in control, PS-deprived and PS-recovery animals. With the head-restrained preparation, we showed that the LPGi contains neurons specifically active during PS (PS-On neurons), neurons inactive during PS (PS-Off neurons) and neurons indifferent to the sleep-waking cycle. After injection of CTb in the facial nucleus, the neurons of which are hyperpolarized during PS, the largest population of Fos/CTb neurons visualized in the medulla in the PS-recovery condition was observed in the LPGi. After injection of CTb in the LPGi itself and PS-recovery, the nucleus containing the highest number of Fos/CTb neurons, moreover bilaterally, was the sublaterodorsal nucleus (SLD). The SLD is known as the pontine executive PS area and triggers PS through glutamatergic neurons. We propose that, during PS, the LPGi is strongly excited by the SLD and hyperpolarizes the motoneurons of the facial nucleus in addition to local and locus coeruleus PS-Off neurons, and by this means contributes to PS genesis.

The neuronal network responsible for paradoxical sleep and its dysfunctions causing narcolepsy and rapid eye movement (REM) behavior disorder.

Rapid eye movement (REM) sleep behavior disorder (RBD) is a parasomnia characterized by the loss of muscle atonia during paradoxical (REM) sleep (PS). Conversely, cataplexy, one of the key symptoms of narcolepsy, is a striking sudden episode of muscle weakness triggered by emotions during wakefulness, and comparable to REM sleep atonia. The neuronal dysfunctions responsible for RBD and cataplexy are not known. In the present review, we present the most recent results on the neuronal network responsible for PS. Based on these results, we propose an updated integrated model of the mechanisms responsible for PS and explore different hypotheses explaining RBD and cataplexy. We propose that RBD is due to a specific degeneration of a sub-population of PS-on glutamatergic neurons specifically responsible of muscle atonia, localized in the caudal pontine sublaterodorsal tegmental nucleus (SLD). Another possibility is the occurrence in RBD patients of a specific lesion of the glycinergic/GABAergic pre-motoneurons localized in the medullary ventral gigantocellular reticular nucleus. Conversely, cataplexy in narcoleptics would be due to the activation during waking of the caudal PS-on SLD neurons responsible for muscle atonia. A phasic glutamatergic excitatory pathway from the central amygdala to the SLD PS-on neurons activated during emotion would induce such activation. In normal conditions, the glutamate excitation would be blocked by the simultaneous excitation by the hypocretins of the PS-off GABAergic neurons localized in the ventrolateral periaqueductal gray and the adjacent deep mesencephalic reticular nucleus, gating the activation of the PS-on SLD neurons.

Novel experience induces persistent sleep-dependent plasticity in the cortex but not in the hippocampus.

Episodic and spatial memories engage the hippocampus during acquisition but migrate to the cerebral cortex over time. We have recently proposed that the interplay between slow-wave (SWS) and rapid eye movement (REM) sleep propagates recent synaptic changes from the hippocampus to the cortex. To test this theory, we jointly assessed extracellular neuronal activity, local field potentials (LFP), and expression levels of plasticity-related immediate-early genes (IEG) arc and zif-268 in rats exposed to novel spatio-tactile experience. Post-experience firing rate increases were strongest in SWS and lasted much longer in the cortex (hours) than in the hippocampus (minutes). During REM sleep, firing rates showed strong temporal dependence across brain areas: cortical activation during experience predicted hippocampal activity in the first post-experience hour, while hippocampal activation during experience predicted cortical activity in the third post-experience hour. Four hours after experience, IEG expression was specifically upregulated during REM sleep in the cortex, but not in the hippocampus. Arc gene expression in the cortex was proportional to LFP amplitude in the spindle-range (10-14 Hz) but not to firing rates, as expected from signals more related to dendritic input than to somatic output. The results indicate that hippocampo-cortical activation during waking is followed by multiple waves of cortical plasticity as full sleep cycles recur. The absence of equivalent changes in the hippocampus may explain its mnemonic disengagement over time.

Fast modulation of prefrontal cortex activity by basal forebrain noncholinergic neuronal ensembles.

Traditionally, most basal forebrain (BF) functions have been attributed to its cholinergic neurons. However, the majority of cortical-projecting BF neurons are noncholinergic and their in vivo functions remain unclear. We investigated how BF modulates cortical dynamics by simultaneously recording

Localization of the GABAergic and non-GABAergic neurons projecting to the sublaterodorsal nucleus and potentially gating paradoxical sleep onset.

We recently determined in rats that iontophoretic application of bicuculline or gabazine [two GABAa antagonists] and kainic acid (a glutamate agonist) in the sublaterodorsal nucleus (SLD) induces with a very short latency a paradoxical sleep-like state. From these results, we proposed that GABAergic and glutamatergic inputs to the SLD paradoxical sleep (PS)-executive neurons gate the onset of PS [R. Boissard et al. (2002) Eur. J. Neurosci., 16, 1959-1973]. We therefore decided to determine the origin of the GABAergic and non-GABAergic inputs to the SLD combining ejection of a retrograde tracer [cholera-toxin B subunit (CTb)] with glutamate decarboxylase (GAD) immunohistochemistry. The presence of GAD-immunoreactive neurons in the SLD was confirmed. Then, following CTb ejections centred on the SLD, combined with GAD and CTb immunohistochemistry, double-labelled cells were observed in the mesencephalic and pontine reticular nuclei and to a lesser extent the parvicellular reticular nucleus. A large number of GAD-negative retrogradely labelled cells was also seen in these structures as well as in the primary motor area of the frontal cortex, the central nucleus of the amygdala, the ventral and lateral bed nucleus of the stria terminalis, the lateral hypothalamic area, the lateral and ventrolateral periaqueductal grey and the lateral paragigantocellular reticular nucleus. From these results, we propose that the activation of PS-executive neurons from the SLD is due to the removal of a tonic inhibition from GABAergic neurons localized in the SLD, and the mesencephalic and pontine reticular nuclei. Strong non-GABAergic inputs to the SLD could be excitatory and responsible for the tonic glutamatergic input on the PS-on neurons we have previously described. They could also terminate on SLD GABAergic interneurons and be indirectly responsible for the inhibition of the PS-on neurons during waking and slow-wave sleep.

The rat ponto-medullary network responsible for paradoxical sleep onset and maintenance: a combined microinjection and functional neuroanatomical study.

The neuronal network responsible for paradoxical sleep (PS) onset and maintenance has not previously been identified in the rat, unlike the cat. To fill this gap, this study has developed a new technique involving the recording of sleep-wake states in unanaesthetized head-restrained rats whilst locally administering pharmacological agents by microiontophoresis from glass multibarrel micropipettes, into the dorsal pontine tegmentum and combining this with functional neuroanatomy. Pharmacological agents used for iontophoretic administration included carbachol, kainic acid, bicuculline and gabazine. The injection sites and their efferents were then identified by injections of anterograde (phaseolus vulgaris leucoagglutinin) or retrograde (cholera toxin B subunit) tracers through an adjacent barrel of the micropipette assembly and by C-Fos immunostaining. Bicuculline, gabazine and kainic acid ejections specifically into the pontine sublaterodorsal nucleus (SLD) induced within a few minutes a PS-like state characterized by a continuous muscle atonia, low voltage EEG and a lack of reaction to stimuli. In contrast, carbachol ejections into the SLD induced wakefulness. In PHA-L, glycine and C-Fos multiple double-labelling experiments, anterogradely labelled fibres originating from the SLD were seen apposed on glycine and C-Fos positive neurons (labelled after 90 min of pharmacologically induced PS-like state) from the ventral gigantocellular and parvicellular reticular nuclei. Altogether, these data indicate that the SLD nuclei contain a population of neurons playing a crucial role in PS onset and maintenance. Furthermore, they suggest that GABAergic disinhibition and glutamate excitation of these neurons might also play a crucial role in the onset of PS.