Ralph Lydic

Evolving concepts of sleep cycle generation: from brain centers to neuronal populations

As neurophysiological investigations of sleep cycle control have provided an increasingly detailed picture of events at the cellular level, the concept that the sleep cycle is generated by the interaction of multiple, anatomically distributed sets of neurons has gradually replaced the hypothesis that sleep is generated by a single, highly localized neuronal oscillator. Cell groups that discharge during rapid-eye-movement (REM) sleep (REM-on) and neurons that slow or cease firing during REM sleep (REM-off) have long been thought to comprise at least two neurochemically distinct populations. The fact that putatively cholinoceptive and/or cholinergic (REM-on) and putatively aminergic (REM-off) cell populations discharge reciprocally over the sleep cycle suggests a causal interdependence. In some brain stem areas these cell groups are not anatomically segregated and may instead be neurochemically mixed (interpenetrated). This finding raises important theoretical and practical issues not anticipated in the original reciprocal-interaction model. The electrophysiological evidence concerning the REM-on and REM-off cell groups suggests a gradient of sleep-dependent membrane excitability changes that may be a function of the connectivity strength within an anatomically distributed neuronal network. The connectivity strength may be influenced by the degree of neurochemical interpenetration between the REM-on and REM-offcells. Recognition of these complexities forces us to revise the reciprocal-interaction model and to seek new methods to test its tenets. Cholinergic microinjection experiments indicate that some populations of REM-on cells can execute specific portions of the REM sleep syndrome or block the generation of REM sleep. This observation suggests that the order of activation within the anatomically distributed generator populations may be critical in determining behavioral outcome. Support for the cholinergic tenets of the reciprocal-interaction model has been reinforced by observations from sleep-disorders medicine. Specific predictions of the reciprocal-interaction model and suggestions for testing these predictions are enumerated for future experimental programs that aim to understand the cellular and molecular basis of the mammalian sleep cycle.

Pontogeniculooccipital waves: spontaneous visual system activity during rapid eye movement sleep

Summary1.Pontogeniculooccipital (PGO) waves are recorded during rapid eye movement (REM) sleep from the pontine reticular formation, lateral geniculate bodies, and occipital cortex of many species.2.PGO waves are associated with increased visual system excitability but arise spontaneously and not via stimulation of the primary visual afferents. Both auditory and somatosensory stimuli influence PGO wave activity.3.Studies using a variety of techniques suggest that the pontine brain stem is the site of PGO wave generation. Immediately prior to the appearance of PGO waves, neurons located in the region of the brachium conjunctivum exhibit bursts of increased firing, while neurons in the dorsal raphe nuclei show a cessation of firing.4.The administration of pharmacological agents antagonizing noradrenergic or serotonergic neurotransmission increases the occurrence of PGO waves independent of REM sleep. Cholinomimetic administration increases the occurrence of both PGO waves and other components of REM sleep.5.Regarding function, the PGO wave-generating network has been postulated to inform the visual system about eye movements, to promote brain development, and to facilitate the response to novel environmental stimuli.

The time-course of dorsal raphe discharge, PGO waves, and muscle tone averaged across multiple sleep cycles

Long-term recordings of dorsal raphe (DRN) activity were obtained from cats chronically implanted with microwires. The continuous time-course of DRN discharge, PGO waves, and muscle tone was quantified across multiple sleep cycles. DRN activity profiles were inversely correlated with PGO waves, biphasically related to muscle tone, and varied with sleep cycle phase. The role of DRN as a putative regulator of behavioral state and/or specific physiological variables is discussed.

Fentanyl and morphine, but not remifentanil, inhibit acetylcholine release in pontine regions modulating arousal.

BACKGROUNDnOpioids inhibit the rapid eye movement (REM) phase of sleep and decrease acetylcholine (ACh) release in medial pontine reticular formation (mPRF) regions contributing to REM sleep generation. It is not known whether opioids decrease ACh release by acting on cholinergic cell bodies or on cholinergic axon terminals. This study used in vivo microdialysis to test the hypothesis that opioids decrease ACh levels at cholinergic neurons in the laterodorsal tegmental nuclei (LDT) and LDT axon terminals in the mPRF.nnnMETHODSnNine male cats were anesthetized with halothane, and ACh levels within the mPRF or LDT were assayed using microdialysis and high-pressure liquid chromatography (HPLC). ACh levels were analyzed in response to dialysis of the mPRF and LDT with Ringers solution (control), followed by dialysis with Ringers solution containing morphine sulfate (MSO4) or naloxone. ACh in the mPRF also was measured during either dialysis delivery or intravenous infusion of remifentanil and during dialysis delivery of fentanyl.nnnRESULTSnCompared with dialysis of Ringers solution, microdialysis with MSO4 decreased ACh by 23% in the mPRF and by 30% in the LDT. This significant decrease in ACh was antagonized by naloxone. MSO4 and fentanyl each caused a dose-dependent decrease in mPRF ACh when delivered by dialysis. Remifentanil delivered by continuous intravenous infusion or by dialysis into the mPRF did not alter mPRF ACh.nnnCONCLUSIONSnMorphine inhibits ACh at the cholinergic cell body region (LDT) and the terminal field in the mPRF. ACh in the mPRF was not altered by remifentanil and was significantly decreased by fentanyl. Thus, MSO4 and fentanyl disrupt cholinergic neurotransmission in the LDT-mPRF network known to modulate REM sleep and cortical electroencephalographic activation. These data are consistent with the possibility that inhibition of pontine cholinergic neurotransmission contributes to arousal state disruption by opioids.

Pontine Cholinergic Mechanisms Modulate the Cortical Electroencephalographic Spindles of Halothane Anesthesia

Background Halothane anesthesia causes spindles in the electroencephalogram (EEG), but the cellular and molecular mechanisms generating these spindles remain incompletely understood. The current study tested the hypothesis that halothane-induced EEG spindles are regulated, in part, by pontine cholinergic mechanisms. Methods Adult male cats were implanted with EEG electrodes and trained to sleep in the laboratory. Approximately 1 month after surgery, animals were anesthetized with halothane and a microdialysis probe was stereotaxically placed in the medial pontine reticular formation (mPRF). Simultaneous measurements were made of mPRF acetylcholine release and number of cortical EEG spindles during halothane anesthesia and subsequent wakefulness. In additional experiments, carbachol (88 mM) was microinjected into the mPRF before halothane anesthesia to determine whether enhanced cholinergic neurotransmission in the mPRF would block the ability of halothane to induce cortical EEG spindles. Results During wakefulness, mPRF acetylcholine release averaged 0.43 pmol/10 min of dialysis. Halothane at 1 minimum alveolar concentration decreased acetylcholine release (0.25 pmol/10 min) while significantly increasing the number of cortical EEG spindles. Cortical EEG spindles caused by 1 minimum alveolar concentration halothane were not significantly different in waveform, amplitude, or number from the EEG spindles of nonrapid eye movement sleep. Microinjection of carbachol into the mPRF before halothane administration caused a significant reduction in number of halothane-induced EEG spindles. Conclusions Laterodorsal and pedunculopontine tegmental neurons, which provide cholinergic input to the mPRF, play a causal role in generating the EEG spindles of halothane anesthesia.

Human brain contains vasopressin and vasoactive intestinal polypeptide neuronal subpopulations in the suprachiasmatic region

The suprachiasmatic nuclei (SCN) and retinohypothalamic tract ( RHT ) in the anterior hypothalamus have been postulated to play an important role in the timing of daily biological rhythms in mammals. Although physiological studies have described circadian rhythms in man, the presence of an RHT or SCN has not been conclusively demonstrated in the human brain. Immunocytochemical identification of distinct ventral vasoactive intestinal polypeptide (VIP) containing and dorsal vasopressin containing neuronal subpopulations in the human suprachiasmatic region provides correlative evidence of neuronal clusters which are homologous to discrete cell groups in the SCN of other mammalian species. Manipulation of the circadian system has been used to treat some affective illnesses and other physiological timing disorders. Characterization of the neural substrates underlying human circadian rhythms could be useful in the development of future treatment modalities and is essential for understanding normal human circadian organization.

Role of the suprachiasmatic nuclei in the circadian timing system of the squirrel monkey. I. The generation of rhythmicity

The circadian organization of squirrel monkey (Saimiri sciureus) drinking behavior was evaluated before and after the placement of radiofrequency lesions which completely destroyed the suprachiasmatic nuclei (SCN) in 4 monkeys and partially ablated the SCN in another 4 animals. In continuous illumination (LL: 600 lux) prior to surgery, each monkey had a precise free-running circadian rhythm of drinking behavior with a period of 25.31 +/- 0.21 h (means +/- S.E.M.). By 4-6 weeks following the lesions, the temporal organization of drinking behavior had become disrupted, but a statistically significant free-running circadian rhythm was still detectable by time series analyses. Subsequently, the circadian organization of drinking behavior in 7 out of 8 monkeys gradually decayed with either no statistically significant rhythmicity or only weak circadian and/or ultradian rhythmicity detected by time series analyses by 10-92 weeks post-lesion. The remaining animal which maintained a statistically reliable free-running rhythm in drinking behavior received the least damage (less than 50%) to the SCN. Despite the major alterations in the temporal patterning of behavior, the overall amount of drinking behavior per 24 h was unchanged. The SCN are thus essential for maintaining the circadian organization of squirrel monkey drinking behavior. However, the existence of residual circadian rhythmicity following SCN lesions and the gradual decay of circadian organization thereafter suggest that the SCN may coordinate the activity of circadian oscillators which lie outside its borders.

Sleep Disruption and Increased Apneas after Pontine Microinjection of Morphine

The medial pontine reticular formation (mPRF) is a cholinoceptive brain stem region known to play a key role in regulating rapid eye movement (REM) sleep and state-dependent ventilatory depression. Numerous lines of evidence have shown that opioids inhibit both cholinergic neurotransmission and REM sleep. The present study examined the hypothesis that morphine applied to the cholinoceptive mPRF would inhibit REM sleep and alter ventilation. In six cats, guide cannulas were chronically implanted to permit pontine microinjection of morphine sulfate, naloxone, and the cholinergic agonist carbachol. After each mPRF microinjection, 2-h polygraphic recordings quantified respiratory frequency and the percent of time spent in states of wakefulness, non-REM sleep, and REM sleep. The results show that mPRF administration of morphine significantly inhibited REM sleep and that this REM sleep inhibitory effect was blocked by pretreating the mPRF with naloxone. Apneic episodes were increased after injection of morphine alone, and the apneas were decreased by the cholinergic agonist carbachol. The results also demonstrated that the ability of microinjected morphine to inhibit REM sleep was dose-dependent and site-dependent. Considered together, the site-localization, pharmacologic blocking, and dose-response data support the hypothesis that specific regions of the mPRF can contribute to the long-recognized ability of morphine to inhibit REM sleep and alter respiratory control.

Halothane decreases pontine acetylcholine release and increases eeg spindles

This study tested the hypothesis that halothane anesthesia would cause decreased acetylcholine (ACh) release within the medial pontine reticular formation (mPRF). ACh was collected by microdialysis and measured by high pressure liquid chromatography during wakefulness and during halothane-induced anesthesia. The electroencephalogram (EEG) showed that spindles were a reliable indicator of anesthetic depth. There was a statistically significant disease in ACh release during halothane anesthesia compared with ACh release during wakefulness. Spindles always disappeared during noxious stimulation and during emergence from anesthesia when pontine ACh levels began to increase. These results are consistent with previous data concerning brain stem cholinergic influences on thalamocortical spindle generation, and suggest that similar mechanisms generate cortical spindles during natural sleep and halothane anesthesia.

Nitric oxide synthase inhibition decreases pontine acetylcholine release

THIS study tested the hypothesis that inhibition of nitric oxide synthase (NOS) in the medial pontine reticular formation (mPRF) would cause decreased acetylcholine (ACh) release. Microdialysis of cat mPRF permitted measurement of ACh during states of wakefulness, non-rapid eye movement (NREM) sleep and rapid eye movement (REM) sleep. ACh release during microdialysis with Ringers (control) was compared to ACh release during microdialysis with 10 mM NG-nitro-L-arginine (NLA). The NOS inhibitor NLA caused a significant reduction in ACh released from the mPRF during wakefulness, NREM sleep, and REM sleep. This reduction in mPRF ACh release elicited by NLA suggests that nitric oxide (NO) contributes to cholinergic neurotrans-mission in the pontine reticular formation.