Pei-Lu Yi

The Central Serotonergic System Mediates the Analgesic Effect of Electroacupuncture on Zusanli (ST36) Acupoints

Evidence in the past decade indicates that the mechanisms of anti-nociception of electroacupuncture (EAc) involve actions of neuropeptides (i.e., enkephalin and endorphin) and monoamines (i.e., serotonin and norepinephrine) in the central nervous system. Our present results using a subcutaneous injection of formalin to test pain sensation in mice provide further understanding of the involvement of serotonin in the actions of EAc-induced analgesia. Our observations show that (1) EAc at three different frequencies (2, 10 and 100 Hz) elicited an anti-nociceptive effect as determined by behavioral observations of reduced hindpaw licking; (2) exogenously intracerebroventricular administration of 5-hydroxytryptamine (5-HT) exhibited an analgesic effect, which partially mimicked the analgesic actions of EAc; (3) the anti-nociception of EAc at different frequencies was attenuated after reduced biosynthesis of serotonin by the administration of the tryptophan hydroxylase inhibitor, P-chlorophenylalanine, and (4) the 5-HT(1A) and 5-HT(3) receptor antagonists, pindobind-5-HT(1A) and LY-278584, respectively, blocked three different frequencies of EAc-induced analgesic effects, but the anti-nociceptive effect of 100 Hz EAc was potentiated by the 5-HT(2) receptor antagonist, ketanserin. These observations suggest that 5-HT(1A) and 5-HT(3) receptors partially mediate the analgesic effects of EAc, but that the 5-HT(2) receptor is conversely involved in the nociceptive response.

Endogenous Opiates in the Nucleus Tractus Solitarius Mediate Electroacupuncture-Induced Sleep Activities in Rats

Electroacupuncture (EA) possesses various therapeutic effects, including alleviation of pain, reduction of inflammation and improvement of sleep disturbance. The mechanisms of EA on sleep improvement, however, remain to be determined. It has been stated in ancient Chinese literature that the Anmian (EX17) acupoint is one of the trigger points that alleviates insomnia. We previously demonstrated that EA stimulation of Anmian acupoints in rats during the dark period enhances non-rapid eye movement (NREM) sleep, which involves the induction of cholinergic activity in the nucleus tractus solitarius (NTS). In addition to cholinergic activation of the NTS, activation of the endogenous opioidergic system may also be a mechanism by which acupuncture affects sleep. Therefore, this study was designed to investigate the involvement of the NTS opioidergic system in EA-induced alterations in sleep. Our present results indicate that EA of Anmian acupoints increased NREM sleep, but not rapid eye movement sleep, during the dark period in rats. This enhancement in NREM sleep was dose-dependently blocked by microinjection of opioid receptor antagonist, naloxone, and the μ-opioid receptor antagonist, naloxonazine, into the NTS; administrations of δ-receptor antagonist, natrindole, and the κ-receptor antagonist, nor-binaltrophimine, however, did not affect EA-induced alterations in sleep. Furthermore, β-endorphin was significantly increased in both the brainstem and hippocampus after the EA stimuli, an effect blocked by administration of the muscarinic antagonist scopolamine into the NTS. Our findings suggest that mechanisms of EA-induced NREM sleep enhancement may be mediated, in part, by cholinergic activation, stimulation of the opiodergic neurons to increase the concentrations of β-endorphin and the involvement of the μ-opioid receptors.

Electrical stimulation of left anterior thalamic nucleus with high-frequency and low-intensity currents reduces the rate of pilocarpine-induced epilepsy in rats.

PURPOSE Bilateral electrical stimulation of anterior nuclei of thalamus (ANT) has shown promising effects on epileptic seizures. However, bilateral implantation increases the risk of surgical complications and side effects. This study was undertaken to access the effectiveness of a stimulation paradigm involving high frequency and low intensity currents to stimulate the left ANT in rats. METHODS Male Sprague-Dawley rats were implanted with electroencephalogram (EEG) electrodes, and an additional concentric bipolar stimulation electrode into either the left or right ANT. The stimulus was a train of pulses (90 μs duration each) delivered with a frequency of 200 Hz and a current intensity of 50 μA. Thalamic stimuli were started 1 h before the first intraperitoneal pilocarpine injection (i.p., 300 mg/kg), and were applied for 5 h. RESULTS EEG documented seizure activity and status epilepticus (SE) developed in 87.5% of rats treated with no ANT stimulation after a single dose of pilocarpine. Left ANT stimulation significantly increased the tolerance threshold for pilocarpine-induced EEG seizure activity; 20% of rats developed their EEG documented seizure activity after receiving the first dose, whereas 50%, 10% and 20% of rats did not develop seizure activity until they had received the 2nd, 3rd and 4th pilocarpine injection at 1-h intervals. Moreover, left thalamic stimulation reduced the occurrences of both EEG documented seizure activity and SE induced by single-dose pilocarpine to 25%. However, our result demonstrated that little effect on the occurrence rate of seizures and SE was found when rats received right ANT stimulation. CONCLUSIONS These results suggest that continuously 5-h left ANT stimulation with high frequency and low intensity currents, beginning from 1h before the pilocarpine administration, may successfully reduce the occurrence rate of EEG documented seizure activity and SE development in rats.

Biphasic effects of baicalin, an active constituent of Scutellaria baicalensis Georgi, in the spontaneous sleep-wake regulation.

AIM OF THE STUDY Baicalin is an active compound originating from the root of Scutellaria baicalensis Georgi, which has been used for anti-inflammation, anti-bacteria, anti-hypertension, anti-allergy and sedation since ancient China, though the neuronal mechanisms involved in the sedative effect is still unclear. Baicalin possesses the ability to decrease the expression of pro-inflammatory cytokines and nuclear factor (NF)-κB activity. Furthermore, baicalin has demonstrated an anxiolytic-like effect via activation of γ-aminobutyric acid-A (GABA(A)) receptors. Pro-inflammatory cytokines (e.g. interleukin (IL)-1, IL-6, and tumor necrosis factor (TNF)-α) and the GABAergic system promote sleep. This study was designed to determine whether the GABA(A) receptor activation and/or the suppression of pro-inflammatory cytokines mediate(s) baicalin-induced sleep alterations. MATERIALS AND METHODS Baicalin was intracerebroventricularly (ICV) administered 20 min either prior to the beginning of the light period or before the onset of the dark period. Electroencephalogram (EEG) and gross body movement were acquired for sleep analysis. Pharmacological blockade of IL-1 and GABA(A) receptors were employed to elucidate the involvements of IL-1 and GABA(A) receptors in baicalin-induced sleep alterations. IL-1β concentrations obtained after baicalin administration in several distinct brain regions were determined by ELISA. RESULTS ICV administration of baicalin decreased slow wave sleep (SWS) during the first 2h of the light period. Rapid eye movement sleep (REMS) was not altered. The blockade of IL-1β-induced SWS enhancement by baicalin suggests that the antagonism of IL-1 receptors is involved in baicalin-induced SWS decrement during the light period. However, IL-1β concentrations during the light period were not altered after baicalin administration. In contrast, baicalin increased both SWS and REMS during hours 8-10 of the dark (active) period when baicalin was administered at the beginning of the dark period, and its effects were blocked by the GABA(A) receptor antagonist bicuculline. CONCLUSION Baicalin exhibits biphasic effects on sleep-wake regulation; the decrease of SWS during the light period and increases of SWS and REMS during the dark period. Inhibition of IL-1 action and enhancement of GABA(A) receptor activity may mediate baicalins effects during the light and dark period, respectively.

TNF-NF-κB signaling mediates excessive somnolence in hemiparkinsonian rats

Daytime somnolence is common in patients with Parkinsons disease (PD); however there is a lack of understanding of the cellular mechanisms involved in mediating these effects. It has been hypothesized that microglial activation and the subsequent increase of pro-inflammatory cytokines play an important role in the pathogenesis of PD. Because some cytokines are involved in the regulation of sleep, this study was designed to determine if tumor necrosis factor (TNF) and interleukin-1beta (IL-1beta), mediate daytime somnolence in the proteasome inhibitor (MG-132)-induced hemiparkinsonian rat model. Our results indicated that microglial activation caused the loss of dopaminergic neurons in the substantia nigra, and the expression of TNF-alpha, but not IL-1beta, increased in the midbrain and hypothalamus in MG-132-induced hemiparkinsonian rats. Slow-wave sleep (SWS) increased after the induction of hemiparkinsonism, but rapid eye movement (REM) sleep was not consistently altered. Application of the TNF receptor fragment (TNFRF) blocked hemiparkinsonism-induced SWS alteration, whereas the IL-1 receptor antagonist (IL-1ra) exhibited no effect. Increased nuclear translocation of NF-kappaB in the midbrain, and the blockade of SWS enhancement in MG-132-induced hemiparkinsonian rats by an inhibitor of NF-kappaB activation indicate that the TNF-NF-kappaB cascade is a critical mediator of MG-132 hemiparkinsonian-induced sleep alteration. This observation suggests potential therapeutic interventions to target the excessive daytime somnolence in patients with PD.

Occurrence of epilepsy at different zeitgeber times alters sleep homeostasis differently in rats.

STUDY OBJECTIVES Controversial sleep disruptions (e.g., poor nighttime sleep and daytime somnolence) are common in epilepsy patients. Sleep is known to be regulated by homeostatic factors, which mediate sleep propensity, and the circadian oscillator, a clocklike mechanism. However, it is unknown how epileptic episodes that occur at different zeitgeber times (ZTs) alter sleep regulation. This study was designed to elucidate the sleep disruptions associated with epilepsy and their underlying mechanisms by delivering kindled epilepsy at different ZTs: ZT0, ZT6, and ZT13. DESIGN Kindled epilepsy was induced at 3 different ZTs, and sleep-wake activities were analyzed before and after full-blown seizure. Ribonuclease protection assay, radioimmunoassay, and immunohistochemistry were respectively employed to determine the levels of interleukin-1 mRNA, corticosterone, and PER1 protein. SETTING The experiments were performed at Neurophysiology Laboratory at National Taiwan University. PARTICIPANT AND INTERVENTIONS: Male Sprague-Dawley rats were implanted with electroencephalograph (EEG) electrodes, a bipolar stimulating electrode, and a guide cannula. Kindling stimuli were delivered via a bipolar electrode placed in the right central nucleus of the amygdala. MEASUREMENT AND RESULTS Kindled epilepsy occurring at ZT0 and ZT13 predominantly affected homeostatic factors, whereas ZT6-kindling stimuli altered the circadian oscillator. ZT0-kindling decreased rapid eye movement (REM) and non-REM (NREM) sleep, which was mediated by corticotrophin-releasing hormone, but did not alter the rhythm of sleep-wake fluctuation. On the other hand, ZT13-kindling enhanced interleukin-1 and consequently increased NREM sleep without altering the sleep-wake fluctuation. Nevertheless, the expression of PER1 protein in suprachiasmatic nucleus of the hypothalamus and the circadian rhythm of sleep fluctuation were respectively advanced 6 h and 2 h when kindling stimulation was delivered at ZT6. Shifts of sleep circadian rhythm and PER1 oscillation induced by ZT6-kindling were blocked by administration of hypocretin receptor antagonist SB334867 into the SCN, indicating the involvement of hypocretin. CONCLUSION These observations suggest that the occurrence of epilepsy at different ZTs alters sleep processes differently. CITATION Yi PL; Chen YJ; Lin CT; Chang FC. Occurrence of epilepsy at different zeitgeber times alters sleep homeostasis differently in rats. SLEEP 2012;35(12):1651-1665.

The involvement of metabotropic glutamate receptors in long-term depression of N-methyl-D-aspartate receptor-mediated synaptic potential in the rat hippocampus.

The frequency-dependent long-term modifications of pharmacologically isolated N-methyl-D-aspartate (NMDA) receptor-mediated excitatory postsynaptic potential (EPSPNMDA) was studied. Intracellular recordings were obtained from CA1 cells of rat hippocampal slices and in the presence of 6-cyano-7-nitroquinoxaline-2,3-dione (10 microM) and bicuculline (20 microM) which block non-NMDA and GABAA receptors, respectively. Low-frequency stimulation at 5 Hz resulted in a long-term depression (LTD) of EPSPNMDA in 12 of 17 cells. However, when the stimulus frequency was increased to 30 Hz, a long-term potentiation (LTP) of EPSPNMDA was observed in 7 out of 9 cells. The LTD was not affected by pretreating the slices with okadaic acid (0.5-1 microM) suggesting that activation of endogenous protein phosphatase is not responsible for this process. In the presence of L-2-amino-3-phosphonopropionic acid (50 microM) or (RS)-alpha-methyl-4-carboxyphenylglycine (200 microM), 5 Hz tetanization resulted in LTP instead of LTD. These results suggest that activation of metabotropic glutamate receptor (mGluR) is necessary for the induction of EPSPNMDA LTD and blockade of mGluR unmasks a LTP.

Kappa-opioid receptors in the caudal nucleus tractus solitarius mediate 100 hz electroacupuncture-induced sleep activities in rats.

Previous results demonstrated that 10 Hz electroacupuncture (EA) of Anmian acupoints in rats during the dark period enhances slow wave sleep (SWS), which involves the induction of cholinergic activity in the caudal nucleus tractus solitarius (NTS) and subsequent activation of opioidergic neurons and μ-receptors. Studies have shown that different kinds of endogenous opiate peptides and receptors may mediate the consequences of EA with different frequencies. Herein, we further elucidated that high-frequency (100 Hz)-EA of Anmian enhanced SWS during the dark period but exhibited no direct effect on rapid eye movement (REM) sleep. High-frequency EA-induced SWS enhancement was dose-dependently blocked by microinjection of naloxone or κ-receptor antagonist (nor-binaltorphimine) into the caudal NTS, but was affected neither by μ- (naloxonazine) nor δ-receptor antagonists (natatrindole), suggesting the role of NTS κ-receptors in the high-frequency EA-induced SWS enhancement. Current and previous results depict the opioid mechanisms of EA-induced sleep.

Activation of GABAergic pathway by hypocretin in the median raphe nucleus (MRN) mediates stress-induced theta rhythm in rats.

The frequency of electroencephalograms (EEGs) is predominant in theta rhythm during stress (e.g., footshock) in rats. Median raphe nucleus (MRN) desynchronizes hippocampal theta waves via activation of GABAergic neurons in the medial septum-diagonal band of Broca (MS-DBB), a theta rhythm pacemaker. Increased hypocretin mediates stress responses in addition to the maintenance of wakefulness. Hypocretin receptors are abundant in the MRN, suggesting a possible role of hypocretin in modulating stress-induced theta rhythm. Our results indicated that the intensity of theta waves was enhanced by footshock and that a hypocretin receptor antagonist (TCS1102) suppressed the footshock-induced theta waves. Administration of hypocretin-1 (1 and 10 μg) and hypocretin-2 (10 μg) directly into the MRN simulated the effect of footshock and significantly increased theta waves. Co-administration of GABA(A) receptor antagonist, bicuculline, into the MRN blocked the increase of theta waves induced by hypocretins or footshock. These results suggested that stress enhances the release of hypocretins, activates GABAergic neurons in the MRN, blocks the ability of MRN to desynchronize theta waves, and subsequently increases the intensity of theta rhythm.

Activation of metabotropic glutamate receptors in conjunction with postsynaptic depolarization triggers a long-term depression of the N-methyl-D-aspartate receptor-mediated synaptic potential in the rat hippocampus

The mechanism responsible for long-term depression (LTD) of pharmacologically isolated N-methyl-D-aspartate (NMDA) receptor-mediated excitatory postsynaptic potential (EPSP(NMDA)) was studied. Intracellular recordings were made from CA1 cells of rat hippocampal slices in the presence of 6-cyano-7-nitroquinoxaline-2,3-dione (10 &mgr;M) and picrotoxin (50 &mgr;M), which block non-NMDA and GABA(A) receptors, respectively. Intracellular injections of depolarizing pulses (500 ms, 0.3-0.7 nA) at 1 Hz for 5 min in the absence of synaptic stimulation caused a persistent increase in the amplitude of EPSP(NMDA). However, coupling postsynaptic depolarization with synaptic activity induced LTD. The EPSP(NMDA) LTD could be blocked by L-2-amino-3-phosphonopropionic acid (50 &mgr;M) or (RS)-alpha-methyl-4-carboxyphenylglycine (200 &mgr;M), specific antagonists for metabotropic glutamate receptors (mGluR). Furthermore, application of trans-1-aminocyclopentane-1,3-dicarboxylic acid (t-ACPD, 50 &mgr;M), a specific mGluR agonist, in conjunction with postsynaptic depolarizing elicited LTD. In contrast, the mGluR agonists quisqualate or t-ACPD when given alone produced a sustained enhancement of EPSP(NMDA). Finally, coupled depolarization did not evoke LTD in slices pretreated with the protein kinase C (PKC) inhibitor calphostin c (60 nM). The present results demonstrate that activation of mGluR is necessary for the induction of LTD of EPSP(NMDA) and suggest that NMDA receptors are subject to bidirectional regulation by mGluR. Furthermore, the induction of LTD is likely to involve the stimulation of PKC. Copyright 1995 S. Karger AG, Basel