Impact of ovariectomy and CO2 inhalation on microglia morphology in select brainstem and hypothalamic areas regulating breathing in female rats.

The neural network that regulates breathing shows a significant sexual dimorphism. Ovarian hormones contribute to this distinction as, in rats, ovariectomy reduces the ventilatory response to CO2. Microglia are neuroimmune cells that are sensitive to neuroendocrine changes in their environment. When reacting to challenging conditions, these cells show changes in their morphology that reflect an augmented capacity for producing pro- and anti-inflammatory cytokines. Based on evidence suggesting that microglia contribute to sex-based differences in reflexive responses to hypercapnia, we hypothesized that ovariectomy and hypercapnia promote microglial reactivity in selected brain areas that regulate breathing. We used ionized calcium-binding-adapter molecule-1 (Iba1) immunolabeling to compare the density and morphology of microglia in the locus coeruleus (LC), the caudal medullary raphe, the caudal part of the nucleus of the tractus solitarius (cNTS), and the paraventricular nucleus of the hypothalamus (PVN). Tissue was obtained from SHAM (metaestrus) female rats or following ovariectomy. Rats were exposed to normocapnia or hypercapnia (5% CO2, 20 min). Ovariectomy and hypercapnia did not affect microglial density in any of the structures studied. Ovariectomy promoted a reactive phenotype in the cNTS and LC, as indicated by a larger morphological index. In these structures, hypercapnia had a relatively modest opposing effect; the medullary raphe or the PVN were not affected. We conclude that ovarian hormones attenuate microglial reactivity in CO2/H+ sensing structures. These data suggest that microglia may contribute to neurological diseases in which anomalies of respiratory control are associated with cyclic fluctuations of ovarian hormones or menopause.

Central deficiency of norepinephrine synthesis and norepinephrinergic neurotransmission contributes to seizure-induced respiratory arrest.

Sudden unexpected death in epilepsy (SUDEP) is the leading cause of mortality in patients with intractable epilepsy. However, the pathogenesis of SUDEP seems to be poorly understood. Our previous findings showed that the incidence of seizure-induced respiratory arrest (S-IRA) was markedly reduced by atomoxetine in a murine SUDEP model. Because the central norepinephrine α-1 receptor (NEα-1R) plays a vital role in regulating respiratory function, we hypothesized that the suppression of S-IRA by atomoxetine was mediated by NE/NEα-1R interactions that can be reversed by NEα-1R antagonism. We examined whether atomoxetine-mediated suppression of S-IRA evoked by either acoustic stimulation or pentylenetetrazole (PTZ) in DBA/1 mice can be reversed by intraperitoneal (IP) and intracerebroventricular (ICV) administration of prazosin, a selective antagonist of NEα-1R. The content and activity of tyrosine hydroxylase (TH), a rate-limiting enzyme for NE synthesis, in the lower brainstem was measured by ELISA. Electroencephalograms (EEG) were obtained from using the PTZ-evoked SUDEP model. In our models, atomoxetine-mediated suppression of S-IRA evoked by either acoustic stimulation or PTZ was significantly reversed by low doses of IP and ICV prazosin. Neither repetitive acoustic stimulation nor S-IRA reduced TH levels in lower brainstem. However, the enzyme activity of TH levels in lower brainstem was significantly increased by mechanical ventilation with DBA/1 mice, which makes the dying DBA/1 mice suffering from S-IRA and SUDEP recover. EEG data showed that although the protective effect of atomoxetine was reversed by prazosin, neither drug suppressed EEG activity. These data suggest that deficient synthesis of NE and norepinephrinergic neurotransmission contributed to S-IRA and that the NEα-1R is a potential therapeutic target for the prevention of SUDEP.

Melatonin attenuates renal sympathetic overactivity and reactive oxygen species in the brain in neurogenic hypertension.

Sympathetic overactivation contributes to the pathogenesis of both experimental and human hypertension. We have previously reported that oxidative stress in sympathetic premotor neurons leads to arterial baroreflex dysfunction and increased sympathetic drive to the kidneys in an experimental model of neurogenic hypertension. In this study, we hypothesized that melatonin, a potent antioxidant, may be protective in the brainstem regions involved in the tonic and reflex control of blood pressure (BP) in renovascular hypertensive rats. Neurogenic hypertension was induced by placing a silver clip (gap of 0.2 mm) around the left renal artery, and after 5 weeks of renal clip placement, the rats were treated orally with melatonin (30 mg/kg/day) by gavage for 15 days. At the end of melatonin treatment, we evaluated baseline mean arterial pressure (MAP), renal sympathetic nerve activity (rSNA), and the baroreflex control of heart rate (HR) and rSNA. Reactive oxygen species (ROS) were detected within the brainstem regions by dihydroethidium staining. Melatonin treatment effectively reduced baseline MAP and sympathoexcitation to the ischemic kidney in renovascular hypertensive rats. The baroreflex control of HR and rSNA were improved after melatonin treatment in the hypertensive group. Moreover, there was a preferential decrease in ROS within the rostral ventrolateral medulla (RVLM) and the nucleus of the solitary tract (NTS). Therefore, our study indicates that melatonin is effective in reducing renal sympathetic overactivity associated with decreased ROS in brainstem regions that regulate BP in an experimental model of neurogenic hypertension.

Anti-Migraine Effect of the Herbal Combination of Chuanxiong Rhizoma and Cyperi Rhizoma and UPLC-MS/MS Method for the Simultaneous Quantification of the Active Constituents in Rat Serum and Cerebral Cortex.

Chuanxiong Rhizoma and Cyperi Rhizoma (CRCR), an ancient and classic formula comprised of Chuanxiong Rhizoma and Cyperi Rhizoma in a weight ratio of 1:2, has long been used for curing migraine. This study aimed to explore their anti-migraine effect and active constituents. A nitroglycerin (NTG)-induced migraine model in rats was established to evaluate pharmacological effects. Cerebral blood flow was detected by a laser Doppler perfusion monitor. The levels of endothelin-1 (ET-1), γ-aminobutyric acid (GABA), nitric oxide synthase (NOS), nitric oxide (NO), 5-hydroxytryptamine (5-HT), 5-hydoxyindoleacetic acid (5-HIAA), calcitonin gene-related peptide (CGRP) and ß-endorphin (ß-EP) were quantified with enzyme-linked immunosorbent assay. CGRP and c-Fos mRNA expression were quantified with quantitative real-time polymerase chain reaction. A UPLC-MS/MS method was developed and validated for the simultaneous quantification of active constituents in rat serum and cerebral cortex. CRCR significantly increased cerebral blood flow, decreased the levels of ET-1, GABA and NOS, and increased the levels of 5-HT, 5-HIAA and ß-EP in NTG-induced migraine rats. CGRP levels and CGRP mRNA expression, as well as c-Fos mRNA expression in the brainstem were markedly down-regulated with the treatment of CRCR. After oral administration of CRCR, ferulic acid (FA), senkyunolide A (SA), 3-n-butylphthalide (NBP), Z-ligustilide (LIG), Z-3-butylidenephthalide (BDPH), cyperotundone (CYT), nookatone (NKT) and α-cyperone (CYP) were qualified in rat serum and cerebral cortex. The above results suggested that CRCR showed powerfully therapeutic effects on migraine via increasing the cerebral blood flow, decreasing the expression of CGRP and c-Fos mRNA, and regulating the releasing of ET-1, GABA, NOS, 5-HT, 5-HIAA, CGRP and ß-EP in the serum and brainstem, consequently relieving neurogenic inflammation. The active constituents in CRCR for treating migraine were FA, SA, NBP, LIG, BDPH, CYT, NKT and CYP. These findings contributed for the further use of CRCR as a combinational and complementary phytomedicine for migraine treatment.

Chronic Consumption of Fructose Induces Behavioral Alterations by Increasing Orexin and Dopamine Levels in the Rat Brain.

It has been widely described that chronic intake of fructose causes metabolic alterations which can be associated with brain function impairment. In this study, we evaluated the effects of fructose intake on the sleep⁻wake cycle, locomotion, and neurochemical parameters in Wistar rats. The experimental group was fed with 10% fructose in drinking water for five weeks. After treatment, metabolic indicators were quantified in blood. Electroencephalographic recordings were used to evaluate the sleep architecture and the spectral power of frequency bands. Likewise, the locomotor activity and the concentrations of orexin A and monoamines were estimated. Our results show that fructose diet significantly increased the blood levels of glucose, cholesterol, and triglycerides. Fructose modified the sleep⁻wake cycle of rats, increasing the waking duration and conversely decreasing the non-rapid eye movement sleep. Furthermore, these effects were accompanied by increases of the spectral power at different frequency bands. Chronic consumption of fructose caused a slight increase in the locomotor activity as well as an increase of orexin A and dopamine levels in the hypothalamus and brainstem. Specifically, immunoreactivity for orexin A was increased in the ventral tegmental area after the intake of fructose. Our study suggests that fructose induces metabolic changes and stimulates the activity of orexinergic and dopaminergic neurons, which may be responsible for alterations of the sleep⁻wake cycle.

The GABAergic Gudden's dorsal tegmental nucleus: A new relay for serotonergic regulation of sleep-wake behavior in the mouse.

Serotonin (5-HT) neurons are involved in wake promotion and exert a strong inhibitory influence on rapid eye movement (REM) sleep. Such effects have been ascribed, at least in part to the action of 5-HT at post-synaptic 5-HT1A receptors (5-HT1AR) in the brainstem, a major wake/REM sleep regulatory center. However, the neuroanatomical substrate through which 5-HT1AR influence sleep remains elusive. We therefore investigated whether a brainstem structure containing a high density of 5-HT1AR mRNA, the GABAergic Gudden's dorsal tegmental nucleus (DTg), may contribute to 5-HT-mediated regulatory mechanisms of sleep-wake stages. We first found that bilateral lesions of the DTg promote wake at the expense of sleep. In addition, using local microinjections into the DTg in freely moving mice, we showed that local activation of 5-HT1AR by the prototypical agonist 8-OH-DPAT enhances wake and reduces deeply REM sleep duration. The specific involvement of 5-HT1AR in the latter effects was further demonstrated by ex vivo extracellular recordings showing that the selective 5-HT1AR antagonist WAY 100635 prevented DTg neuron inhibition by 8-OH-DPAT. We next found that GABAergic neurons of the ventral DTg exclusively targets glutamatergic neurons of the lateral mammillary nucleus (LM) in the posterior hypothalamus by means of anterograde and retrograde tracing techniques using cre driver mouse lines and a modified rabies virus. Altogether, our findings strongly support the idea that 5-HT-driven enhancement of wake results from 5-HT1AR-mediated inhibition of DTg GABAergic neurons that would in turn disinhibit glutamatergic neurons in the mammillary bodies. We therefore propose a Raphe→DTg→LM pathway as a novel regulatory circuit underlying 5-HT modulation of arousal.

GLP-1 receptor agonist liraglutide exerts central action to induce ß-cell proliferation through medulla to vagal pathway in mice.

Endogenous GLP-1 and GLP-1 receptor agonists (GLP-1RAs) regulate glucose metabolism via common and distinct mechanisms. Postprandial release of GLP-1 is modest and it is degraded by DPP-4 within 2 min, and hence it cannot enter the brain in substantial amount. In contrast, DPP-4-resistant GLP-1RAs are administered at 10 times higher concentration than endogenous GLP-1 level, which enables them to reach several brain regions including ARC and AP, the areas implicated in glucose metabolism. Hence, some of the effects of GLP-1RAs observed clinically and experimentally, including pancreatic ß-cell proliferation, are thought to involve the brain. However, the effects of centrally acting GLP-1/GLP-1RAs on glucose metabolism and underlying neural mechanism are unclear. This study aimed to establish the link of central GLP-1/GLP-1RA action to pancreatic ß-cell proliferation. Both subcutaneous (SC) and intracerebroventricular (ICV) injections of liraglutide increased the number of pancreatic ß-cells expressing Ki67 and PCNA, proliferation markers, in C57BL/6J mice. This effect was induced by single ICV administration of liraglutide at relatively low dose that was incapable of suppressing food intake. These SC and ICV liraglutide-induced effects were inhibited by 50% and 70%, respectively, by pretreatment with atropine, a muscarinic receptor blocker. ICV liraglutide induced c-Fos expression in the area postrema (AP), nucleus tractus solitaries (NTS), and dorsal motor nucleus of the vagus (DMX) of the brain stem. These results demonstrate that central action of liraglutide induces pancreatic ß-cell proliferation via the pathway involving the brain stem AP/NTS/DMX area and vagus nerve. This route is highly sensitive to GLP-1/GLP-1RA. Hence, this brain-pancreatic ß-cell pathway may operate in type 2 diabetic patients treated with GLP-RAs and serve to counteract the reduction of ß-cell mass.

Time-dependent effects of olanzapine treatment on the expression of histidine decarboxylase, H1 and H3 receptor in the rat brain: The roles in olanzapine-induced obesity.

Antipsychotic treatment, particularly olanzapine and clozapine, induces severe obesity. The Histamine H1 receptor is considered to be an important contributor to olanzapine-induced obesity, however how olanzapine modulates the histaminergic system is not sufficiently understood. This study examined the effect of olanzapine on key molecules of the histaminergic system, including histidine decarboxylase (HDC), H1 receptor (H1R) and H3 receptor (H3R), in the brain at different stages of olanzapine-induced obesity. During short-term treatment (8-day), olanzapine increased hypothalamic HDC mRNA expression and H1R binding in the arcuate nucleus (Arc) and ventromedial hypothalamus (VMH), without changing H3R binding density. HDC mRNA and Arc H1R binding were positively correlated with increased food intake, feeding efficiency and weight gain. When the treatment was extended to 16 and 36 days, H1R binding was increased not only in the hypothalamic Arc and VMH but also in the brainstem dorsal vagal complex (DVC). The H1R bindings in the Arc, VMH and DVC were positively correlated with weight gain induced by olanzapine treatment. However, the expression of HDC and H3R mRNA was not increased. These results suggest that olanzapine time-dependently modulates histamine neurotransmission, which suggested the different neuronal mechanisms underlying different stages of weight gain development. Treatment targeting the H1R may be effective for both short- and long-term olanzapine-induced weight gain.

Curcumin decreases astrocytic reaction after gliotoxic injury in the rat brainstem.

Recent studies have demonstrated that curcumin (Cur) has antioxidant, anti-inflammatory and anti-fibrotic effects. Ethidium bromide (EB) injections into the central nervous system (CNS) are known to induce local oligodendroglial and astrocytic loss, resulting in primary demyelination and neuroinflammation. Peripheral astrogliosis is seen around the injury site with increased immunoreactivity to glial fibrillary acidic protein (GFAP). This investigation aimed to evaluate the effect of Cur administration on astrocytic response following gliotoxic injury. Wistar rats were injected with EB into the cisterna pontis and treated, or not, with Cur (100 mg/kg/day, intraperitoneal route) during the experimental period. Brainstem sections were collected at 15, 21 and 31 days after EB injection and processed for GFAP immunohistochemical staining. Astrocytic reactivity was measured in a computerized system for image analysis. In Cur-treated rats, the GFAP-stained area around the lesion was significantly smaller in all periods after EB injection compared to untreated animals, showing that Cur reduces glial scar development following injury.

Curcumin decreases astrocytic reaction after gliotoxic injury in the rat brainstem / Curcumina reduz a reação astrocitária após injúria gliotóxica no tronco encefálico de ratos

ABSTRACT Recent studies have demonstrated that curcumin (Cur) has antioxidant, anti-inflammatory and anti-fibrotic effects. Ethidium bromide (EB) injections into the central nervous system (CNS) are known to induce local oligodendroglial and astrocytic loss, resulting in primary demyelination and neuroinflammation. Peripheral astrogliosis is seen around the injury site with increased immunoreactivity to glial fibrillary acidic protein (GFAP). This investigation aimed to evaluate the effect of Cur administration on astrocytic response following gliotoxic injury. Wistar rats were injected with EB into the cisterna pontis and treated, or not, with Cur (100 mg/kg/day, intraperitoneal route) during the experimental period. Brainstem sections were collected at 15, 21 and 31 days after EB injection and processed for GFAP immunohistochemical staining. Astrocytic reactivity was measured in a computerized system for image analysis. In Cur-treated rats, the GFAP-stained area around the lesion was significantly smaller in all periods after EB injection compared to untreated animals, showing that Cur reduces glial scar development following injury.

RESUMO Estudos recentes têm demonstrado que a curcumina (Cur) possui efeitos antioxidantes, anti-inflamatórios e antifibróticos. Sabe-se que a injeção de brometo de etídio (EB) no sistema nervoso central induz a perda oligodendroglial e astrocitária, resultando em desmielinização primária e neuroinflamação. Astrogliose periférica é observada ao redor da lesão com aumento da imunorreatividade à proteína glial fibrilar ácida (GFAP). A presente investigação objetivou avaliar o efeito da Cur sobre a resposta astrocitária após injúria gliotóxica. Ratos Wistar foram injetados com EB na cisterna basal e tratados ou não com Cur (100 mg/kg/dia, via intraperitoneal) durante o período experimental. Amostras do tronco encefálico foram coletadas aos 15, 21 e 31 dias pós-injeção de EB e processadas para estudo imuno-histoquímico para a GFAP. A reatividade astrocitária foi medida em um sistema computadorizado para análise de imagem. Nos ratos tratados com Cur, a área marcada para GFAP foi significantemente menor em todos os períodos pós-injeção de EB, indicando que a Cur reduz o desenvolvimento da cicatriz glial após injúria.

Mortality with brainstem seizures from focal 4-aminopyridine-induced recurrent hippocampal seizures.

OBJECTIVE: Sudden unexplained death in epilepsy is the leading cause of death in young adult epilepsy patients, typically occurring during the early postictal period, presumably resulting from brainstem and cardiorespiratory dysfunction. We hypothesized that ictal discharges in the brainstem disrupt the cardiorespiratory network, causing mortality. To study this hypothesis, we chose an animal model comprising focal unilateral hippocampal injection of 4-aminopyridine (4-AP), which produced focal recurrent hippocampal seizures with secondary generalization in awake, behaving rats. METHODS: We studied ictal and interictal intracranial electrographic activity (iEEG) in 23 rats implanted with a custom electrode array into the hippocampus, the contralateral cortex, and brainstem. The hippocampal electrodes contained a cannula to administer the potassium channel blocker and convulsant (4-AP). iEEG was recorded continuously before, during, and after seizures induced by 4-AP infusion into the hippocampus. RESULTS: The control group (n = 5) was monitored for 2-3 months, and the weekly baseline iEEG recordings showed long-term stability. The low-dose group (1 µL 4-AP, 40 mm, n = 5) exhibited local electrographic seizures without spread to the contralateral cerebral cortex or brainstem. The high-dose group (5 µL 4-AP, 40 mm, n = 3) had several hippocampal electrographic seizures, which spread contralaterally and triggered brainstem discharges within 40 min, and were associated with violent motor seizures followed by dyspnea and respiratory arrest, with cortical and hippocampal iEEG flattening. The group that received high-dose 4-AP without brainstem implantation (n = 5) had similar seizure-related respiratory difficulties. Finally, five rats that received high-dose 4-AP without EEG recording also developed violent motor seizures with postictal respiratory arrest. Following visualized respiratory arrest in groups III, IV, and V, manual respiratory resuscitation was successful in five of 13 animals. SIGNIFICANCE: These studies show that hippocampal seizure activity can spread or trigger brainstem epileptiform discharges that may cause mortality, possibly mediated by respiratory network dysfunction.

Bottom-Up and Top-Down Mechanisms of General Anesthetics Modulate Different Dimensions of Consciousness.

There has been controversy regarding the precise mechanisms of anesthetic-induced unconsciousness, with two salient approaches that have emerged within systems neuroscience. One prominent approach is the "bottom up" paradigm, which argues that anesthetics suppress consciousness by modulating sleep-wake nuclei and neural circuits in the brainstem and diencephalon that have evolved to control arousal states. Another approach is the "top-down" paradigm, which argues that anesthetics suppress consciousness by modulating the cortical and thalamocortical circuits involved in the integration of neural information. In this article, we synthesize these approaches by mapping bottom-up and top-down mechanisms of general anesthetics to two distinct but inter-related dimensions of consciousness: level and content. We show how this explains certain empirical observations regarding the diversity of anesthetic drug effects. We conclude with a more nuanced discussion of how levels and contents of consciousness interact to generate subjective experience and what this implies for the mechanisms of anesthetic-induced unconsciousness.

Role of the area postrema in the hypophagic effects of oleoylethanolamide.

The satiety-promoting action of oleoylethanolamide (OEA) has been associated to the indirect activation of selected brain areas, such as the nucleus of the solitary tract (NST) in the brainstem and the tuberomammillary (TMN) and paraventricular (PVN) nuclei in the hypothalamus, where noradrenergic, histaminergic and oxytocinergic neurons play a necessary role. Visceral ascending fibers were hypothesized to mediate such effects. However, our previous findings demonstrated that the hypophagic action of peripherally administered OEA does not require intact vagal afferents and is associated to a strong activation of the area postrema (AP). Therefore, we hypothesized that OEA may exert its central effects through the direct activation of this circumventricular organ. To test this hypothesis, we subjected rats to the surgical ablation of the AP (APX rats) and evaluated the effects of OEA (10mgkg-1 i.p.) on food intake, Fos expression, hypothalamic oxytocin (OXY) immunoreactivity and on the expression of dopamine beta hydroxylase (DBH) in the brainstem and hypothalamus. We found that the AP lesion completely prevented OEA's behavioral and neurochemical effects in the brainstem and the hypothalamus. Moreover OEA increased DBH expression in AP and NST neurons of SHAM rats while the effect in the NST was absent in APX rats, thus suggesting the possible involvement of noradrenergic AP neurons. These results support the hypothesis of a necessary role of the AP in mediating OEA's central effects that sustain its pro-satiety action.

Frontal-Brainstem Pathways Mediating Placebo Effects on Social Rejection.

Placebo treatments can strongly affect clinical outcomes, but research on how they shape other life experiences and emotional well-being is in its infancy. We used fMRI in humans to examine placebo effects on a particularly impactful life experience, social pain elicited by a recent romantic rejection. We compared these effects with placebo effects on physical (heat) pain, which are thought to depend on pathways connecting prefrontal cortex and periaqueductal gray (PAG). Placebo treatment, compared with control, reduced both social and physical pain, and increased activity in the dorsolateral prefrontal cortex (dlPFC) in both modalities. Placebo further altered the relationship between affect and both dlPFC and PAG activity during social pain, and effects on behavior were mediated by a pathway connecting dlPFC to the PAG, building on recent work implicating opioidergic PAG activity in the regulation of social pain. These findings suggest that placebo treatments reduce emotional distress by altering affective representations in frontal-brainstem systems.SIGNIFICANCE STATEMENT Placebo effects are improvements due to expectations and the socio-medical context in which treatment takes place. Whereas they have been extensively studied in the context of somatic conditions such as pain, much less is known of how treatment expectations shape the emotional experience of other important stressors and life events. Here, we use brain imaging to show that placebo treatment reduces the painful feelings associated with a recent romantic rejection by recruiting a prefrontal-brainstem network and by shifting the relationship between brain activity and affect. Our findings suggest that this brain network may be important for nonspecific treatment effects across a wide range of therapeutic approaches and mental health conditions.

Effect of the Level of Anesthesia on the Auditory Brainstem Response in the Emei Music Frog (Babina daunchina).

Anesthesia is known to affect the auditory brainstem response (ABR) in mice, rats, birds and lizards. The present study investigated how the level of anesthesia affects ABR recordings in an amphibian species, Babina daunchina. To do this, we compared ABRs evoked by tone pip stimuli recorded from 35 frogs when Tricaine methane sulphonate (MS-222) anesthetic immersion times varied from 0, 5 and 10 minutes after anesthesia induction at sound frequencies between 0.5 and 6 kHz. ABR thresholds increased significantly with immersion time across the 0.5 kHz to 2.5 kHz frequency range, which is the most sensitive frequency range for hearing and the main frequency range of male calls. There were no significant differences for anesthetic levels across the 3 kHz to 6 kHz range. ABR latency was significantly longer in the 10 min group than in the 0 and 5 min groups at frequencies of 0.5, 1.0, 1.5, 2.5 kHz, while ABR latency did not differ across the 3 kHz to 4 kHz range and at 2.0 kHz. Taken together, these results show that the level of anesthesia affects the amplitude, threshold and latency of ABRs in frogs.

Spreading depolarization triggered by elevated potassium is weak or absent in the rodent lower brain.

We examined in live coronal slices from rat and mouse which brain regions generate potassium-triggered spreading depolarization (SDKt). This technique simulates cortical spreading depression, which underlies migraine aura in the intact brain. An SDKt episode was evoked by increasing bath [K+]o and recorded as a propagating front of elevated light transmittance representing transient neuronal swelling in gray matter of neocortex, hippocampus, striatum, and thalamus. In contrast, SDKt was not imaged in hypothalamic nuclei or brainstem with exception of those nuclei near the dorsal brainstem surface. In rat slices, single neurons were whole-cell current clamped during SDKt. "Higher" neurons depolarized to near zero millivolts indicating SDKt generation. In contrast, seven types of neurons in hypothalamus and brainstem only slowly depolarized without generating SDKt, supporting our imaging findings. Therefore, SDKt is not a default of CNS neurons but rather displays a region-specific susceptibility, similar to anoxic depolarization, which we have proposed is correlated with a region's vulnerability to traumatic brain injury. In the higher brain, SDKt may be a vestigial spreading depolarization that originally evolved to shut down and vasoconstrict gray matter regions more exposed to impact and contusion.

Pharmacodynamic action and mechanism of Du Liang soft capsule, a traditional Chinese medicine capsule, on treating nitroglycerin-induced migraine.

ETHNOPHARMACOLOGICAL RELEVANCE: Du Liang soft capsule (DL) is a traditional Chinese medicine for treating migraines; it is made from two Chinese herbs, including LigusticumstriatumDC., root; Angelica dahurica (Hoffm.) Benth. & Hook.f. ex Franch. & Sav., root. AIM OF THE STUDY: In the present study, we aimed to elucidate the pharmacodynamic action of DL and its mechanism in an animal model of migraines induced by glyceryl trinitrate (GTN). MATERIALS AND METHODS: Sixty rats were randomly divided into six groups, including a normal control group, model control group, positive group (Sumatriptan 0.006gkg-1), and three DL groups (0.44, 1.31 and 3.93gkg-1). All rats were intragastrically treated with the corresponding treatment for 7 consecutive days, and they were subcutaneously injected with GTN (10mgkg-1) 30min after the last treatment, except in the normal control group. After model establishment, the behaviors of all rats, including head scratching, cage climbing, and the development of red ears were observed continuously by digital camera every 30min for 3h. Four hours after GTN treatment, all rats were anaesthetized and the blood and tissue samples were collected. Plasma calcitonin gene related to peptide (CGRP) and endothelin (ET) levels were measured using the radioimmunoassay method, and serum NO was determined by the colorimetric method. Afterwards, the brainstem tissues were dissected and washed with physiological saline, and divided evenly into two parts. One part was used to test the monoamine levels, including levels of 5-hydroxytryptamine (5-HT), norepinephrine (NE) and dopamine (DA), by the fluorometric method, and the other part was used to determine the nuclear factor kappaB (NF-κB) p65, nuclear c-fos, inducible nitric oxide synthase (iNOS), interleukin (IL)-1ß (IL-1ß), and cyclooxygenase-2 (COX-2) levels by Western blot analysis. RESULTS: In the pharmacodynamic action assay, DL (1.31 and 3.93gkg-1) greatly improved the abnormal behaviors of migraine rats, including head scratching and cage climbing, and the development of red ears. In the mechanism assay, compared with the control group, the plasma CGRP and serum NO levels and the brainstem 5-HT, NE and DA levels in the DL administration groups were significantly decreased; and the plasma ET levels were remarkably increased. Moreover, down-regulation of NF-κB p65, c-fos and pro-inflammatory cytokines, including iNOS, IL-1ß and COX-2 in the brainstem in the DL administration groups were observed by Western blot analysis. CONCLUSIONS: The above results suggested that DL has a therapeutic effect on migraines, and its mechanism may be related to adjusting the level of neurotransmitters and vasoactive substances, consequently relieving neurogenic inflammation.