Md. Noor Alam

Sleep-waking discharge patterns of neurons recorded in the rat perifornical lateral hypothalamic area

The perifornical lateral hypothalamic area (PF‐LHA) has been implicated in the control of several waking behaviours, including feeding, motor activity and arousal. Several cell types are located in the PF‐LHA, including projection neurons that contain the hypocretin peptides (also known as orexins). Recent findings suggest that hypocretin neurons are involved in sleep‐wake regulation. Loss of hypocretin neurons in the human disorder narcolepsy is associated with excessive somnolence, cataplexy and increased propensity for rapid eye movement (REM) sleep. However, the relationship of PF‐LHA neuronal activity to different arousal states is unknown. We recorded neuronal activity in the PF‐LHA of rats during natural sleep and waking. Neuronal discharge rates were calculated during active waking (waking accompanied by movement), quiet waking, non‐REM sleep and REM sleep. Fifty‐six of 106 neurons (53 %) were classified as wake/REM‐related. These neurons exhibited peak discharge rates during waking and REM sleep and reduced discharge rates during non‐REM sleep. Wake‐related neurons (38 %) exhibited reduced discharge rates during both non‐REM and REM sleep when compared to that during waking. Wake‐related neurons exhibited significantly higher discharge rates during active waking than during quiet waking. The discharge of wake‐related neurons was positively correlated with muscle activity across all sleep‐waking states. Recording sites were located within the hypocretin‐immunoreactive neuronal field of the PF‐LHA. Although the neurotransmitter phenotype of recorded cells was not determined, the prevalence of neurons with wake‐related discharge patterns is consistent with the hypothesis that the PF‐LHA participates in the regulation of arousal, muscle activity and sleep‐waking states.

Adenosinergic modulation of rat basal forebrain neurons during sleep and waking: neuronal recording with microdialysis

1 The cholinergic system of the basal forebrain (BF) is hypothesized to play an important role in behavioural and electrocortical arousal. Adenosine has been proposed as a sleep‐promoting substance that induces sleep by inhibiting cholinergic neurons of the BF and brainstem. However, adenosinergic influences on the activity of BF neurons in naturally awake and sleeping animals have not been demonstrated. 2 We recorded the sleep‐wake discharge profile of BF neurons and simultaneously assessed adenosinergic influences on wake‐ and sleep‐related activity of these neurons by delivering adenosinergic agents adjacent to the recorded neurons with a microdialysis probe. Discharge rates of BF neurons were recorded through two to three sleep‐wake episodes during baseline (artificial cerebrospinal fluid perfusion), and after delivering an adenosine transport inhibitor (s‐(p‐nitrobenzyl)‐6‐thioinosine; NBTI), or exogenous adenosine, or a selective adenosine A1 receptor antagonist (8‐cyclopentyl‐1,3‐dimethylxanthine; CPDX). 3 NBTI and adenosine decreased the discharge rate of BF neurons during both waking and non‐rapid eye movement (NREM) sleep. In contrast, CPDX increased the discharge rate of BF neurons during both waking and NREM sleep. These results suggest that in naturally awake and sleeping animals, adenosine exerts tonic inhibitory influences on BF neurons, supporting the hypothesized role of adenosine in sleep regulation. 4 However, in the presence of exogenous adenosine, NBTI or CPDX, BF neurons retained their wake‐ and sleep‐related discharge patterns, i.e. still exhibited changes in discharge rate during transitions between waking and NREM sleep. This suggests that other neurotransmitters/neuromodulators also contribute to the sleep‐wake discharge modulation of BF neurons.

GABA-mediated control of hypocretin- but not melanin-concentrating hormone-immunoreactive neurones during sleep in rats

The perifornical‐lateral hypothalamic area (PF‐LHA) has been implicated in the regulation of behavioural arousal. The PF‐LHA contains several cell types including neurones expressing the peptides, hypocretin (HCRT; also called orexin) and melanin‐concentrating hormone (MCH). Evidence suggests that most of the PF‐LHA neurones, including HCRT neurones, are active during waking and quiescent during non‐rapid eye movement (non‐NREM) sleep. The PF‐LHA contains local GABAergic interneurones and also receives GABAergic inputs from sleep‐promoting regions in the preoptic area of the hypothalamus. We hypothesized that increased GABA‐mediated inhibition within PF‐LHA contributes to the suppression of neuronal activity during non‐REM sleep. EEG and EMG activity of rats were monitored for 2 h during microdialytic delivery of artificial cerebrospinal fluid (aCSF) or bicuculline, a GABAA receptor antagonist, into the PF‐LHA in spontaneously sleeping rats during the lights‐on period. At the end of aCSF or bicuculline perfusion, rats were killed and c‐Fos immunoreactivity (Fos‐IR) in HCRT, MCH and other PF‐LHA neurones was quantified. In response to bicuculline perfusion into the PF‐LHA, rats exhibited a dose‐dependent decrease in non‐REM and REM sleep time and an increase in time awake. The number of HCRT, MCH and non‐HCRT/non‐MCH neurones exhibiting Fos‐IR adjacent to the microdialysis probe also increased dose‐dependently in response to bicuculline. However, significantly fewer MCH neurones exhibited Fos‐IR in response to bicuculline as compared to HCRT and other PF‐LHA neurones. These results support the hypothesis that PF‐LHA neurones, including HCRT neurones, are subject to increased endogenous GABAergic inhibition during sleep. In contrast, MCH neurones appear to be subject to weaker GABAergic control during sleep.

Different types of norepinephrinergic receptors are involved in preoptic area mediated independent modulation of sleep-wakefulness and body temperature

The preoptic area is known to regulate sleep-wakefulness and body temperature. It was suggested earlier that though sleep-wakefulness and body temperature may affect each other, the preoptic area mediated influence on those two physiological phenomena is likely to be independent of alteration in each other. Since intrapreoptic area norepinephrine could modulate both those functions, study of that system was undertaken. It was hypothesized that since the preoptic area has different types of norepinephrinergic receptors (viz. alpha 1, alpha 2 and beta), independent modulation of those two functions was probably due to activation or inactivation of separate receptors. Hence, the effects of different agonist and antagonist of those receptors individually as well as in combination into the preoptic area were studied on those two functions in freely moving rats. The results suggest that norepinephrine induced preoptic area mediated influence on the body temperature is primarily regulated by the alpha 1 receptors while the sleep and wakefulness are regulated by alpha 2 and beta receptors, respectively. The finding should help in explaining several poorly understood observations reported earlier and it suggests that similar phenomena may possibly exist in other system involving other neurotransmitters as well.

Interleukin-1β modulates state-dependent discharge activity of preoptic area and basal forebrain neurons: role in sleep regulation

Interleukin‐1β (IL‐1) is a pro‐inflammatory cytokine that has been implicated in the regulation of nonrapid eye movement (nonREM) sleep. IL‐1, IL‐1 receptors and the IL‐1 receptor antagonist (ra) are present normally in discrete brain regions, including the preoptic area (POA) of the hypothalamus and the adjoining magnocellular basal forebrain (BF). The POA/BF have been implicated in the regulation of sleep–wakefulness. We hypothesized that IL‐1 promotes nonREM sleep, in part by altering the state‐dependent discharge activity of POA/BF neurons. We recorded the sleep–wake discharge profiles of 83 neurons in the lateral POA/BF and assessed the effects of IL‐1, IL‐1ra, and IL‐ra + IL‐1 delivered through a microdialysis probe on state‐dependent neuronal discharge activity. IL‐1 decreased the discharge rate of POA/BF neurons as a group (n = 55) but wake‐related and sleep‐related neurons responded differently. IL‐1 significantly decreased the discharge rate of wake‐related neurons. Of 24 wake‐related neurons studied, 19 (79%) neurons exhibited a greater than 20% change in their discharge in the presence of IL‐1 during waking. IL‐1 suppressed the discharge activity of 18 of 19 responsive neurons. Of 13 sleep‐related neurons studied, IL‐1 increased the discharge activity of five and suppressed the discharge activity of four neurons. IL‐1ra increased the discharge activity of four of nine neurons and significantly attenuated IL‐1‐induced effects on neuronal activity of POA/BF neurons (n = 19). These results suggest that the sleep‐promoting effects of IL‐1 may be mediated, in part, via the suppression of wake‐related neurons and the activation of a subpopulation of sleep‐related neurons in the POA/BF.

The Median Preoptic Nucleus Reciprocally Modulates Activity of Arousal-Related and Sleep-Related Neurons in the Perifornical Lateral Hypothalamus

The perifornical–lateral hypothalamic area (PF/LH) contains neuronal groups playing an important role in control of waking and sleep. Among the brain regions that regulate behavioral states, one of the strongest sources of projections to the PF/LH is the median preoptic nucleus (MnPN) containing a sleep-active neuronal population. To evaluate the role of MnPN afferents in the control of PF/LH neuronal activity, we studied the responses of PF/LH cells to electrical stimulation or local chemical manipulation of the MnPN in freely moving rats. Single-pulse electrical stimulation evoked responses in 79% of recorded PF/LH neurons. No cells were activated antidromically. Direct and indirect transynaptic effects depended on sleep–wake discharge pattern of PF/LH cells. The majority of arousal-related neurons, that is, cells discharging at maximal rates during active waking (AW) or during AW and rapid eye movement (REM) sleep, exhibited exclusively or initially inhibitory responses to stimulation. Sleep-related neurons, the cells with elevated discharge during non-REM and REM sleep or selectively active in REM sleep, exhibited exclusively or initially excitatory responses. Activation of the MnPN via microdialytic application of l-glutamate or bicuculline resulted in reduced discharge of arousal-related and in excitation of sleep-related PF/LH neurons. Deactivation of the MnPN with muscimol caused opposite effects. The results indicate that the MnPN contains subset(s) of neurons, which exert inhibitory control over arousal-related and excitatory control over sleep-related PF/LH neurons. We hypothesize that MnPN sleep-active neuronal group has both inhibitory and excitatory outputs that participate in the inhibitory control of arousal-promoting PF/LH mechanisms.

Preoptic/anterior hypothalamic neurons : thermosensitivity in wakefulness and non rapid eye movement sleep

Thermosensitive neurons of the preoptic/anterior hypothalamic area (POAH) have been implicated in the regulation of both body temperature and non rapid eye movement (NREM) sleep. During NREM sleep, a majority of POAH warm-sensitive neurons (WSN) exhibit increased discharge compared to wakefulness. Cold-sensitive neurons (CSN) exhibit reduced discharge in NREM sleep compared to wakefulness. To further study the mechanism underlying these processes, the present study compared discharge rate and thermosensitivity (discharge rate change/degree C) of WSNs and CSNs in NREM sleep and wakefulness in freely moving adult cats. The thermosensitivity of 24 WSNs and 31 CSNs from the medial POAH was determined from responses to local POAH warming and cooling. WSNs with increased discharge in NREM sleep exhibited increased thermosensitivity during NREM sleep compared to wakefulness. CSNs with decreased discharge during NREM sleep exhibited decreased thermosensitivity in NREM sleep. The change in thermosensitivity from wakefulness to NREM sleep was correlated with the change in discharge rate in WSNs but not in CSNs. In addition, 9 of 47 neurons that were thermo-insensitive during wakefulness became warm-sensitive during NREM sleep. Changes in POAH neuronal thermosensitivity could be a component of the mechanism for stabilization of state after state transition.

Differential acute influence of medial and lateral preoptic areas on sleep-wakefulness in freely moving rats.

The role of preoptic area (POA) in sleep-wakefulness and related EEG changes is well established. Anatomically the area is divided into medial (mPOA) and lateral (IPOA) portions having different physiological functions. Knowledge regarding the differential role, if any, of those two areas in sleep and wakefulness was lacking in the literature. Therefore, an attempt was made in this study, to investigate the same systematically. Experiments were conducted during day and night in freely moving rats. Electrophysiological parameters defining sleep and wakefulness were recorded before and after reversible inactivation of those two areas separately by microinjection of a local anaesthetic, marcain. The responses were opposite in nature depending upon the time, day or night, when the anaesthetic was applied. During the day, anaesthetization induced wakefulness while during the night, sleep was precipitated. However, anaesthetization of both the areas though induced similar qualitative response, the degree of the responses differed significantly. The results suggest that the mPOA is more effective in maintaining tonic sleep while the IPOA is more potent in the maintenance of tonic wakefulness in the normal rats. The finding supports and fits well with the existing knowledge.

Thermosensitive neurons of the diagonal band in rats: relation to wakefulness and non-rapid eye movement sleep

The thermosensitivity of the neurons in the diagonal band of Broca (DBB) was studied in 12 freely moving rats by determining responses to local cooling or warming with a water perfused thermode. Of 151 neurons studied, 37 (25%) neurons met the criterion for thermosensitivity including 17 warm-sensitive (WSNs) and 20 cold-sensitive neurons (CSNs). The spontaneous discharge rates of WSNs and CSNs were recorded through 1-3 sleep-waking cycles. The discharge of WSNs and CSNs during waking and non-rapid eye movement (NREM) sleep were different. Of 17 WSNs, 10 exhibited increased discharge rates during NREM sleep as compared with waking (NREM/Wake discharge ratio, > 1.2). Of 20 CSNs, 14 discharged more slowly during NREM sleep as compared with waking (NREM/Wake discharge ratio, < 0.8). In both WSNs and CSNs, changes in discharge rate preceded EEG changes at the waking-NREM transition. These results support a hypothesis that the activation of sleep-related WSNs and the deactivation of wake-related CSNs play a role in the onset and regulation of NREM sleep.

Neuronal activity in the preoptic hypothalamus during sleep deprivation and recovery sleep

The preoptic hypothalamus is implicated in sleep regulation. Neurons in the median preoptic nucleus (MnPO) and the ventrolateral preoptic area (VLPO) have been identified as potential sleep regulatory elements. However, the extent to which MnPO and VLPO neurons are activated in response to changing homeostatic sleep regulatory demands is unresolved. To address this question, we continuously recorded the extracellular activity of neurons in the rat MnPO, VLPO and dorsal lateral preoptic area (LPO) during baseline sleep and waking, during 2 h of sleep deprivation (SD) and during 2 h of recovery sleep (RS). Sleep-active neurons in the MnPO (n = 11) and VLPO (n = 13) were activated in response to SD, such that waking discharge rates increased by 95.8 ± 29.5% and 59.4 ± 17.3%, respectively, above waking baseline values. During RS, non-rapid eye movement (REM) sleep discharge rates of MnPO neurons initially increased to 65.6 ± 15.2% above baseline values, then declined to baseline levels in association with decreases in EEG delta power. Increase in non-REM sleep discharge rates in VLPO neurons during RS averaged 40.5 ± 7.6% above baseline. REM-active neurons (n = 16) in the LPO also exhibited increased waking discharge during SD and an increase in non-REM discharge during RS. Infusion of A2A adenosine receptor antagonist into the VLPO attenuated SD-induced increases in neuronal discharge. Populations of LPO wake/REM-active and state-indifferent neurons and dorsal LPO sleep-active neurons were unresponsive to SD. These findings support the hypothesis that sleep-active neurons in the MnPO and VLPO, and REM-active neurons in the LPO, are components of neuronal circuits that mediate homeostatic responses to sustained wakefulness.