1.Morphological study on development of nerve growth factor-positive neurons in the cerebellum of human fetus.
Acta Academiae Medicinae Sinicae 2012;34(1):46-50
OBJECTIVETo investigate the growth and development of nerve growth factor (NGF)-positive neurons in the cerebellum of midanaphase human fetus.
METHODSThe expression of the NGF-positive neurons in the cerebrum of human fetus was observed by immunohistochemical methods, and the integral absorbance (IA) was detected.
RESULTSBy the 3rd to 4th month of gestation, neurons was seen in the ependymal, central, and marginal plate of cerebellum; the nucleus was oval and the neurons had short and small processes. By the 5th to 7th month of gestation, the number of NGF-positive neurons increased, the expressions enhanced, the nucleus was round-, oval-, or fusiform-shaped, the neurons grew larger in size, and the Purkinje cells showed NGF-positive expression. By the 8th to 10th month of gestation, the NGF-positive expression was enhanced with deeper dying, the body of Purkinje cells grew larger gradually, and the number of NGF-positive neurons in the granular cell layer and molecular layer increased. IA of the cerebellar cortical neurons of the 3rd, 4th, 5th, 6th, 7th, and 8th month of gestation showed an increasing trend, and significant difference was observed (P < 0. 05).
CONCLUSIONNGF-positive neurons in the cerebellum play an important role for differentiation, proliferation, migration, and growth of neurons in the cerebellum.
Cerebellum ; cytology ; metabolism ; Fetus ; cytology ; metabolism ; Humans ; Nerve Growth Factor ; metabolism ; Neurons ; cytology ; metabolism ; Purkinje Cells ; metabolism
2.Comparison of electrophysiological properties of vascular smooth muscle cells in different arterioles in guinea pig.
Ke-Tao MA ; Xin-Zhi LI ; Li LI ; Zhi-Ping ZHANG ; Lei ZHAO ; He ZHU ; Jun-Qiang SI
Acta Physiologica Sinica 2010;62(5):421-426
Arterioles are major contributors to the control of systemic blood pressure and local blood flow. In this study, we compared electrophysiological properties of vascular smooth muscle cells (VSMCs) in anterior inferior cerebellar artery (AICA), mesenteric artery (MA) and spiral modiolar artery (SMA) by intracellular microelectrode recording and whole-cell patch clamp recording techniques. Results were shown as below: (1) Intracellular microelectrode recordings were made from VSMCs in AICA, MA and SMA with resting potentials of (-68±1.8) (n=65), (-71±2.4) (n=80) and (-66±2.9) mV (n=58), respectively. There was no significant difference in resting potentials among arterioles. (2) The membrane capacitance and membrane conductance in situ cells were much larger than those in dispersed smooth muscle cells by whole-cell recording techniques, and there was significant difference among arterioles, which were in the order: MA>AICA>SMA. After application of gap junction blocker 2-APB (100 μmol/L), the membrane capacitance and membrane conductance in situ cells were very close with those in single smooth muscle cells. (3) The I/V relation of whole-cell current of dissociated smooth muscle cells (AICA, MA and SMA) showed a prominent outward rectification, and the currents were substantially inhibited by 1 mmol/L 4-AP or 10 mmol/L TEA. When the command voltage was +40 mV, the current densities of VSMCs in AICA, MA and SMA were (26±2.0), (24±1.7) and (18±1.3) pA/pF respectively. SMA showed significant difference in the current density from AICA and MA respectively. These results suggest that the electrophysiological properties of coupling strength of gap junction and current density of smooth muscle cells are different among arterioles in the guinea pig.
Animals
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Arterioles
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cytology
;
physiology
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Cerebellum
;
blood supply
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Cochlea
;
blood supply
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Electrophysiological Phenomena
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Female
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Guinea Pigs
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Male
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Mesenteric Arteries
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cytology
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Muscle, Smooth, Vascular
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cytology
;
physiology
3.Immunohistochemistry of Voltage-Gated Calcium Channel alpha1B Subunit in Mouse Cerebellum.
Seung Hyuk CHUNG ; Kyung Youl LEE ; Ki Hyung KIM ; Chul Tae KIM ; Nam Seob LEE ; Kazuhiko SAWADA ; Hiromi HAGA ; Byung Chul LEE ; Yoshihiro FUKUI ; Man Hee RHEE ; Young Gil JEONG
Journal of Veterinary Science 2002;3(3):175-178
Secretion of neurotransmitters is initiated by voltagegated calcium influx through presynaptic, voltage- gated N-type calcium channels. However, little is known about their cellular distribution in the mouse cerebellum. In the cerebellum, alpha1B immunoreactivity is found mainly on the cell bodies of all Purkinje cells. In addition, the immunoreactivity was detected on a subset of Purkinje cell dendrites, clustered to form a parasagittal array of bands. In the anterior lobe vermis, immunoreactive Purkinje cell dendrites form narrow stripes separated by broad bands of unstained dendrites. Moving caudally through the vermis, these stripes become thicker as a larger fraction of the Purkinje cell dendrites become immunoreactive. This localization study of the alpha1B pore-forming subunits in mouse cerebellum may guide future investigations of the role of calcium channels in neurological pathways.
Animals
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Calcium Channels, N-Type/*metabolism
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Cerebellum/cytology/*metabolism
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Dendrites/metabolism
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Immunohistochemistry
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Mice
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Mice, Inbred BALB C
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Purkinje Cells/metabolism
4.Mitochondrial calcium uniporter protein MCU is involved in oxidative stress-induced cell death.
Yajin LIAO ; Yumin HAO ; Hong CHEN ; Qing HE ; Zengqiang YUAN ; Jinbo CHENG
Protein & Cell 2015;6(6):434-442
Mitochondrial calcium uniporter (MCU) is a conserved Ca(2+) transporter at mitochondrial in eukaryotic cells. However, the role of MCU protein in oxidative stress-induced cell death remains unclear. Here, we showed that ectopically expressed MCU is mitochondrial localized in both HeLa and primary cerebellar granule neurons (CGNs). Knockdown of endogenous MCU decreases mitochondrial Ca(2+) uptake following histamine stimulation and attenuates cell death induced by oxidative stress in both HeLa cells and CGNs. We also found MCU interacts with VDAC1 and mediates VDAC1 overexpression-induced cell death in CGNs. This finding demonstrates that MCU-VDAC1 complex regulates mitochondrial Ca(2+) uptake and oxidative stress-induced apoptosis, which might represent therapeutic targets for oxidative stress related diseases.
Animals
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Apoptosis
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Biological Transport
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Calcium
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metabolism
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Calcium Channels
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metabolism
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Cerebellum
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cytology
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HeLa Cells
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Humans
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Mice
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Mitochondria
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metabolism
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Neurons
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cytology
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metabolism
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Oxidative Stress
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Voltage-Dependent Anion Channels
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metabolism
5.Curcumin alters motor coordination but not total number of Purkinje cells in the cerebellum of adolescent male Wistar rats.
Ginus PARTADIREDJA ; Sutarman ; Taufik Nur YAHYA ; Christiana Tri NURYANA ; Rina SUSILOWATI
Journal of Integrative Medicine 2013;11(1):32-38
OBJECTIVEThe present study aimed at investigating the effects of curcumin on the motor coordination and the estimate of the total number of cerebellar Purkinje cells of adolescent Wistar rats exposed to ethanol.
METHODSThe total of 21 male Wistar rats aged 37 d old were divided into three groups, namely ethanol, ethanol-curcumin, and control groups. The ethanol group received 1.5 g/kg ethanol injected intraperitoneally and water given per oral; the ethanol-curcumin group received 1.5 g/kg ethanol injected intraperitoneally and curcumin extract given per oral; the control group received saline injection and oral water. The treatment was carried out daily for one month, after which the motor coordination performance of the rats was examined using revolving drum apparatus at test days 1, 8, and 15. The rats were finally sacrificed and the cerebellum of the rats was further processed for stereological analysis. The estimate of the total number of Purkinje cells was calculated using physical fractionator method.
RESULTSThe ethanol-curcumin group performed better than both ethanol and control groups in the motor coordination ability at day 8 of testing (P< 0.01). No Purkinje cell loss was observed as a result of one month intraperitoneal injection of ethanol.
CONCLUSIONCurcumin may exert beneficial effects on the motor coordination of adolescent rats exposed to ethanol via undetermined hormetic mechanisms.
Animals ; Cell Count ; Cerebellum ; cytology ; drug effects ; Curcumin ; pharmacology ; Ethanol ; toxicity ; Male ; Motor Activity ; drug effects ; Psychomotor Performance ; Purkinje Cells ; cytology ; drug effects ; Rats ; Rats, Wistar
6.Expression of PSD95 in the Rat Sciatic Nerve.
Hyun Jin YOO ; Ik Hyun CHO ; Jong Hwan LEE ; Nong Hoon CHOE ; Tae Young KANG ; Byung Joon CHANG
Journal of Veterinary Science 2003;4(2):113-116
This study was designed to elucidate the existence of PSD95 in the rat sciatic nerve. Immunohistochemical stains of cryosection and teased fiber of sciatic nerves were performed with goat polyclonal antibody against PSD95. Western blot analysis was also accomplished with the same antibody. We got an interesting result that the rat sciatic nerve obviously showed PSD95 immunoreactivity especially in the nodal and paranodal regions, and we also identified a distinct band of PSD95 by western blot. These results suggest PSD95 exists in the sciatic nerve as well as it does in the central nervous system. We suppose PSD95 may have some important roles in ion channel clustering, junctional plasticity and signal transduction in the peripheral nerves as well.
Animals
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Blotting, Western
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Cerebellum/cytology/metabolism
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Immunohistochemistry
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Nerve Fibers/metabolism/ultrastructure
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Nerve Tissue Proteins/*metabolism
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Rats
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Rats, Sprague-Dawley
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Sciatic Nerve/*metabolism
7.Interleukin-6 prevents cultured cerebellar granule neurons from glutamate-induced neurotoxicity.
Jian-hua LU ; Yi-hua QIU ; Yu-ping PENG
Chinese Journal of Applied Physiology 2006;22(3):310-315
AIMTo explore IL-6 neuroprotection against glutamate-induced neurotoxicity and primary mechanisms involved in this neuroprotection.
METHODSThe cerebellar granule neurons from postnatal 8-day infant rats were chronically exposed to IL-6 for 8 days, and then glutamate stimulated the cultured cerebellar granule neurons for 15 min. Methyl-thiazole-tetrazolium (MTT) assay and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) method were used to observe the changes of neuronal vitality and apoptosis, respectively. Laser scanning confocal microscope (LSCM) and reverse transcription-polymerase chain reaction (RT-PCR) were respectively employed to measure dynamic changes of intracellular Ca2+ levels and expression of gp130 mRNA, a 130-kDa intracellular IL-6 signal-transduction protein, in the neurons.
RESULTSThe chronic IL-6 (2.5, 5 and 10 ng/ml) pretreatment of the cultured cerebellar granule neurons remarkably improved the decreased neuronal vitality by glutamate in a concentration-dependent manner. The neuronal apoptosis induced by glutamate was significantly attenuated by the chronic IL-6 pretreatment. The intracellular Ca2+ overload evoked by glutamate was also inhibited by the chronic IL-6 pretreatment. The expression of gp130 mRNA was dramatically lower in the IL-6-pretreated cerebellar granule neurons than in the IL-6-untreated neurons.
CONCLUSIONIL-6 can protect neurons against glutamate-induced exciting neurotoxicity. The mechanism of IL-6 neuroprotection may be closely related to the suppression of glutamate-induced intracellular Ca2+ overload and mediated by gp130 intracellular signal transduction pathways.
Animals ; Cells, Cultured ; Cerebellum ; cytology ; drug effects ; metabolism ; Glutamic Acid ; toxicity ; Interleukin-6 ; pharmacology ; Neurons ; drug effects ; metabolism ; Neuroprotective Agents ; pharmacology ; Neurotoxicity Syndromes ; metabolism ; Rats ; Rats, Sprague-Dawley
8.Interleukin-6 protects cerebellar granule neurons from NMDA-induced neurotoxicity.
Xiao-Chun WANG ; Yi-Hua QIU ; Yu-Ping PENG
Acta Physiologica Sinica 2007;59(2):150-156
Interleukin-6 (IL-6) is an important cytokine that participates in inflammation reaction and cell growth and differentiation in the immune and nervous systems. However, the neuroprotection of IL-6 against N-methyl-D-aspartate (NMDA)-induced neurotoxicity and the related underlying mechanisms are still not identified. In the present study, the cultured cerebellar granule neurons (CGNs) from postnatal (8-day) infant rats were chronically exposed to IL-6 for 8 d, and then NMDA (100 micromol/L) was applied to the cultured CGNs for 30 min. Methyl-thiazole-tetrazolium (MTT) assay, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) method and confocal laser scanning microscope (CLSM) were used to detect neuronal vitality, apoptosis and dynamic changes of intracellular Ca(2+) levels in the neurons, respectively. Anti-gp130 monoclonal antibody (75 ng/mL) was employed to the cultured CGNs with IL-6 to inhibit IL-6 activity so as to evaluate the role of gp130 (a 130 kDa glucoprotein transducing IL-6 signal) in mediating IL-6 neuroprotection. Western blot was used to measure the expressions of phospho-signal transducer and activator of transcription 3 (STAT3) and phospho-extracellular signal regulated kinase 1/2 (ERK1/2) in the cultured CGNs. The NMDA stimulation of the cultured CGNs without IL-6 pretreatment resulted in a significant reduction of the neuronal vitality, notable enhancement of the neuronal apoptosis and intracellular Ca(2+) overload in the neurons. The NMDA stimulation of the CGNs chronically pretreated with IL-6 caused a remarkable increase in the neuronal vitality, marked suppression of neuronal apoptosis and intracellular Ca(2+) overload in the neurons, compared with that in the control neurons without IL-6 pretreatment. Furthermore, anti-gp130 antibody blocked the inhibitory effect of IL-6 on NMDA-induced intracellular Ca(2+) overload in the neurons. The levels of phospho-STAT3 and phospho-ERK1/2 were significantly higher in IL-6-pretreated CGNs than those in IL-6-untreated neurons. The results suggest that chronic IL-6 pretreatment of CGNs protects the neurons against NMDA-induced neurotoxicity. The neuroprotective effect of IL-6 is closely related to its suppression of NMDA-induced intracellular Ca(2+) overload and is possibly mediated by gp130/JAK-STAT3 and gp130/RAS-ERK1/2 transduction pathways.
Animals
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Animals, Newborn
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Cells, Cultured
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Cerebellum
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cytology
;
drug effects
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metabolism
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Interleukin-6
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physiology
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MAP Kinase Signaling System
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N-Methylaspartate
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antagonists & inhibitors
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toxicity
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Neurons
;
cytology
;
drug effects
;
metabolism
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Neuroprotective Agents
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Rats
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Rats, Sprague-Dawley
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STAT3 Transcription Factor
;
metabolism
9.Protective effect of epigallocatechin-3-gallate on apoptosis of rat cerebellar granule neurons induced by acrylamide.
Chunfang LIU ; Chengmei JIANG ; Lihua ZHOU
Journal of Central South University(Medical Sciences) 2012;37(9):944-950
OBJECTIVE:
To investigate the protective effect of epigallocatechin-3 -gallate (EGCG) on apoptosis of cerebellar granule neurons (CGNs) induced by acrylamide (ACR).
METHODS:
CGNs were cultured with the addition of 5 mmol/L ACR for 24 hours to set up a cell injury model. Prior to ACR treatment, CGNs were treated with different concentrations of EGCG (0, 5, 10, 25, 50, 100 μmol/L) for 48 hours. Neuronal viability was measured with metylthiazdyltetrazolium (MTT). The activity of SOD and the content of MDA were assayed. Hoechst33342 staining was employed to observe morphological changes of the cell nucleus. Reverse transcription-polymerase chain reaction (RT-PCR) was used to measure expression of bcl- 2 mRNA and bax mRNA.
RESULTS:
At the concentrations of 10, 25 or 50 μmol/L, EGCG played a protective role against ACRinduced CGN injury. Compared with ACR injured group (no EGCG), EGCG improved the cell viability, enhanced SOD activity, decreased the level of MDA as well as the cell apoptosis ratio (P<0.05). Bcl-2 mRNA expression was increased and bax mRNA expression was reduced (P<0.05). 25 μmol/L EGCG had the largest effect. However, 100 μmol/L EGCG did not have a significantly protective effect.
CONCLUSION
EGCG at appropriate concentration has protective effect against the CGNs on apoptosis induced by ACR.
Acrylamide
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toxicity
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Animals
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Apoptosis
;
drug effects
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Catechin
;
analogs & derivatives
;
pharmacology
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Cells, Cultured
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Cerebellum
;
cytology
;
drug effects
;
Cytoplasmic Granules
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Female
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Male
;
Neurons
;
cytology
;
drug effects
;
Neuroprotective Agents
;
pharmacology
;
Rats
;
Rats, Sprague-Dawley
10.Acute hypoxia increases outward current and decreases gap junction of VSMCs in guinea-pig anterior inferior cerebellar artery.
Xin-Zhi LI ; Jun-Qiang SI ; Zhong-Shuang ZHANG ; Lei ZHAO ; Li LI ; Ke-Tao MA
Acta Physiologica Sinica 2011;63(6):533-539
The aim of the present study was to investigate the effects of acute hypoxia on the electrophysiological properties of vascular smooth muscle cells (VSMCs) in arteriole. Guinea-pig anterior inferior cerebellar artery (AICA) segments were isolated, and outer layer connective tissue was removed by collagenase A digestion and microforceps. By perfusion with physical saline solution containing no glucose and low oxygen, VSMC model of acute hypoxia was established. The model was studied by whole-cell patch clamp recording technique. Results were shown as below: (1) Acute hypoxia induced an outward current with amplitude of (36.4 ± 9.2) pA at holding potential of -40 mV, and the rest potential (RP) of the VSMCs was hyperpolarized from (-33.2 ± 1.9) mV to (-38.4 ± 1.5) mV. Acute hypoxia increased the outward current of VSMCs in a voltage-dependent manner, this enhancing effect being more pronounced at potentials ranging from 0 to +40 mV. The whole-cell membrane current of VSMCs induced by step command (+40 mV) increased from (650 ± 113) pA to (1 900 ± 197) pA. In the presence of 1 mmol/L tetraethylammonium (TEA), the enhancement of the VSMC membrane current by acute hypoxia was significantly reduced. (2) Acute hypoxia increased the membrane resistance (R(input)) of the VSMCs in AICA from (234 ± 63) MΩ to (1 211 ± 201) MΩ, and decreased the membrane capacitance (C(input)) from (279.3 ± 83.2) pF to (25.4 ± 1.9) pF. In the presence of 30 μmol/L 18β-glycyrrhetinic acid (18βGA) and 10 mmol/L TEA, the effects of acute hypoxia on the membrane current of VSMCs were nearly abolished. These results suggest that acute hypoxia causes vascular hyperpolarization and vasodilation, possibly by activating big conductance Ca(2+)-activated K(+) channels (BK(Ca)) of the VSMCs, and inhibits gap junctions between VSMCs, thus improving microcirculation and localizing the hypoxia-induced damage.
Animals
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Arteries
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physiopathology
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Cerebellum
;
blood supply
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Female
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Gap Junctions
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metabolism
;
physiology
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Guinea Pigs
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Hypoxia
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physiopathology
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In Vitro Techniques
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Male
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Muscle, Smooth, Vascular
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cytology
;
metabolism
;
physiology
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Myocytes, Smooth Muscle
;
metabolism
;
physiology
;
Patch-Clamp Techniques
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Potassium Channels
;
physiology