PURPOSE: To investigate the effects of brimonidine, an alpha-2-adrenergic agonist, on barrier function in ARPE-19 cells by measuring transepithelial resistance (TER). METHODS: ARPE-19 cells were cultured into a confluent monolayer on a microporous filter. Brimonidine was added to the apical medium, and the barrier function of the cells was evaluated by measuring TER. A subset of cells was treated under hypoxic conditions, and the TER changes observed upon administration of brimonidine were compared to those observed in cells in normoxic conditions. RESULTS: The ARPE cell membrane reached a peak resistance of 29.1+/-7.97 Omega cm2 after four weeks of culture. The TER of the cells treated under normoxic conditions increased with brimonidine treatment; however, the TER of the cells treated under hypoxic conditions did not change following the administration of brimonidine. CONCLUSIONS: Barrier function in ARPE-19 cells increased with brimonidine treatment. Understanding the exact mechanism of this barrier function change requires further investigation.
Adrenergic alpha-Agonists/*pharmacology ; Cell Hypoxia/drug effects/physiology ; Cell Line ; Electric Impedance ; Humans ; Quinoxalines/*pharmacology ; Receptors, Adrenergic, alpha-2/*drug effects ; Retinal Pigment Epithelium/*drug effects/*physiology

OBJECTIVETo study the interactions between Ligusticum chuanxiong Hort extract and cardiac muscle membrane receptors.

METHODThe cell membrane of rabbit cardiac muscle was fixed on silicon to make cell membrane stationary phase (CMSP), and then the interactions were studied by comparing the retention characteristics of the extracts from different solvents with those of the antagonists or activators corresponding to known receptors in cardiac muscle membrane, and by competition effect on the retention characteristics of extracts when adding the antagonists or activators into the mobile phase.

RESULTWater extract and ethanol extract both had retentions on CMSP; the retention characteristics of water extract could be affected when water extract was in competition with the antagonists for alpha receptor, and could not be affected when with the activator beta1 receptor.

CONCLUSIONIt is possible that some components in water extract may combine with alpha receptor and no component with beta1 receptor, and that some components in ethanol extract may combine with cardiac muscle cell membrane. The process between active components and receptors in vivo can be imitated through the interactions between drugs and CMSP. The method provides references for the resolution of two applications: to screen the active components from Chinese medicine, and to figure out the type of receptors involved.

Adrenergic alpha-Agonists ; metabolism ; Adrenergic alpha-Antagonists ; metabolism ; Adrenergic beta-Agonists ; metabolism ; Adrenergic beta-Antagonists ; metabolism ; Animals ; Cell Membrane ; metabolism ; Chromatography, High Pressure Liquid ; methods ; Drugs, Chinese Herbal ; isolation & purification ; pharmacology ; Female ; Ligusticum ; chemistry ; Male ; Myocytes, Cardiac ; cytology ; metabolism ; Plants, Medicinal ; chemistry ; Protein Binding ; Rabbits ; Receptors, Adrenergic, alpha ; metabolism ; Receptors, Adrenergic, beta ; metabolism

PURPOSE: Fentanyl was reported to inhibit the alpha1-adrenoceptor agonist-induced contraction. The goal of this in vitro study was to identify the alpha1-adrenoceptor subtype primarily involved in the fentanyl-induced attenuation of phenylephrine-induced contraction in isolated endothelium-denuded rat aorta. MATERIALS AND METHODS: Aortic rings were suspended in order to record isometric tension. Concentration-response curves for phenylephrine (10-9 to 10-5 M) were generated in the presence or absence of one of the following drugs: fentanyl (3x10-7, 10-6, 3x10-6 M), 5-methylurapidil (3x10-8, 10-7, 3x10-7 M), chloroethylclonidine (10-5 M) and BMY 7378 (3x10-9, 10-8, 3x10-8 M). Phenylephrine concentration-response curves were generated in the presence or absence of fentanyl in rings pretreated with either 3x10-9 M prazosin, 10-9 M 5-methylurapidil or 3x10-9 M BMY 7378. RESULTS: Fentanyl (10-6, 3x10-6 M) attenuated phenylephrine-induced contraction in the rat aorta. 5-Methylurapidil and BMY 7378 produced a parallel rightward shift in the phenylephrine concentration-response curve. The pA2 values for 5-methylurapidil and BMY 7378 were estimated to be 7.71 +/- 0.15 and 8.99 +/- 0.24, respectively. Fentanyl (10-6 M) attenuated phenylephrine-induced contraction in rings pretreated with 10-9 M 5-methylurapidil, but did not alter the rings when pretreated with 3x10-9 M BMY 7378. Pretreatment of the rings with chloroethylclonidine showed a 72.9 +/- 2.3% reduction in phenylephrine-induced maximal contraction. CONCLUSION: The results suggest that fentanyl attenuates phenylephrine-induced contraction by inhibiting the pathway involved in the alpha1D-adrenoceptor-mediated contraction of the rat aorta.
Adrenergic alpha-Agonists/*pharmacology ; Adrenergic alpha-Antagonists/*pharmacology ; Animals ; Aorta/*drug effects ; Clonidine/analogs & derivatives/pharmacology ; Fentanyl/*pharmacology ; Male ; Phenylephrine/*pharmacology ; Piperazines/pharmacology ; Rats ; Rats, Sprague-Dawley ; Vasoconstriction/*drug effects

OBJECTIVETo investigate the expression and distribution of alpha1-adrenoceptor subtypes in rabbit submandibular gland and the effect of phenylephrine on salivary secretion.

METHODSThe expressions of alpha1A-, alpha1B-, and alpha1D-adrenoceptor mRNA and protein were investigated by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot in rabbit submandibular gland. Immunohistochemical assay was applied to detect the distribution of alpha1A-, alpha1B-, and alpha1D-adrenoceptor and localization of aquaporin 5 in rabbit submandibular gland. Different concentrations of phenylephrine (1 x 10(-8))-(1 x 10(-6)) mol/L were administrated through a polyethylene tube, which was intubated into Wharton's duct of submandibular gland. Heart rate and blood pressure of rabbits were observed during phenylephrine administration. Salivary flow was measured by the length of moist filter paper (35 mm x 5 mm) within 5 min.

RESULTSAlpha1A-, alpha1B-, and alpha1D-adrenoceptor mRNA and protein were expressed in rabbit submandibular gland. Three alpha1-adrenoceptor subtypes were widely distributed in the membrane and cytoplasma of both acinar and ductal cells. Phenylephrine (1 x 10(-7) mol/L, 100 microl) stimulated effectively salivary secretion without inducing significant alteration of blood pressure and heart rate in rabbit. Immunohistochemical assay showed that aquaporin 5 was mainly localized in the apical and lateral plasma membrane in both acinar and ductal cells in unstimulated condition, while the expression of aquaporin 5 was increased after administration of phenylephrine.

CONCLUSIONSExpression of alpha1A-, alpha1B-, and alpha1D-adrenoceptor mRNA and protein was existed in rabbit submandibular gland. Phenylephrine safely and effectively promoted salivary secretion when it was administrated through Wharton's duct of submandibular gland. The mechanism of phenylephrine on salivary secretion may involve in the increase of expression of aquaporin 5 in the apical and lateral plasma membrane in rabbit submandibular gland. This study will hopefully lead to a novel strategy for clinical treatment of dysfunction of submandibular gland.

Adrenergic alpha-Agonists ; pharmacology ; Animals ; Aquaporin 5 ; metabolism ; Phenylephrine ; pharmacology ; RNA, Messenger ; genetics ; metabolism ; Rabbits ; Receptors, Adrenergic, alpha-1 ; genetics ; metabolism ; Salivation ; drug effects ; Submandibular Gland ; drug effects ; secretion

OBJECTIVE: Dexmedetomidine (DEX), the most specific α ₂-adrenergic receptor agonist widely used for its sedative and analgesic properties, has been reported to upregulate HIF-1α expression to protect hypoxic and ischemic tissues. However, it is largely unclear whether DEX can also upregulate Hypoxia-inducible factor-1 alpha (HIF-1α) expression and its downstream vascular endothelial growth factor-A (VEGFA) in cancer tissues with oxygen-deficient tumor microenvironment. METHODS: We used SMMC-7721 cells, MHCC97-H cells, and a mouse model of orthotopic hepatic carcinoma to explore the effect of DEX on angiogenesis and vasculogenic mimicry (VM) and its mechanism. Under normoxic (20% O ₂) and hypoxic (1% O ₂) conditions, DEX was used to intervene cells, and yohimbine was used to rescue them. RESULTS: The results showed that DEX promoted angiogenesis and VM in human liver cancer cells within a certain dose range, and the addition of yohimbine inhibited this effect. DEX could activate HIF-1α/VEGFA pathway, which was further verified by silencing HIF-1α. Consistently, in vivo results also showed that DEX can up-regulate HIF-1α/VEGFA expression, and enhance the number of VM channels and microvessel density (MVD). CONCLUSION We believe that HIF-1α/VEGFA might be an important signaling pathway by which DEX promotes angiogenesis and VM formation in human hepatocellular carcinoma, whereas α ₂-adrenergic receptor mediation might be the critical mechanisms.
Animals ; Humans ; Mice ; Adrenergic alpha-2 Receptor Agonists/pharmacology* ; Carcinoma, Hepatocellular ; Cardiovascular Physiological Phenomena ; Dexmedetomidine/pharmacology* ; Hypoxia ; Liver Neoplasms/drug therapy* ; Oxygen ; Tumor Microenvironment ; Vascular Endothelial Growth Factor A/genetics* ; Receptors, Adrenergic, alpha-2/metabolism*

The purpose of this study was to investigate the effects of α₂ adrenergic receptor agonist dexmedetomidine on withdrawal symptoms in alcohol-dependent rats and the underlying mechanism, so as to provide a scientific basis for the treatment of alcohol withdrawal syndrome (AWS). Adult Sprague-Dawley (SD) male rats were orally administered with 6% aqueous alcohol continuously for 28 d to establish alcohol drinking model, and then stopped drinking to induce AWS. Enzyme-linked immunosorbent assay (ELISA) was used to determine the content of norepinephrine (NE) in the locus coeruleus and hippocampus of rats. Dexmedetomidine (5, 10, and 20 μg/kg) was intraperitoneally injected respectively when the rats showed significant AWS. In some rats, α₂ adrenergic receptor antagonist yohimbine was injected into hippocampus in advance. The results showed that, compared with the control group, the 6 h withdrawal group exhibited significantly increased AWS score and amount of repeat drinking. The NE contents in hippocampus and locus coeruleus of the last drinking and the 6 h withdrawal groups were significantly increased compared with those of the control group. Dexmedetomidine intervention significantly decreased AWS score and hippocampus NE content in the 6 h withdrawal group, while yohimbine could reverse these effects of dexmedetomidine. These results suggest that dexmedetomidine might improve the withdrawal symptoms in alcohol-dependent rats via activating α₂ adrenergic receptor.
Adrenergic alpha-2 Receptor Agonists/therapeutic use* ; Alcoholism/drug therapy* ; Animals ; Dexmedetomidine/therapeutic use* ; Hippocampus/metabolism* ; Male ; Norepinephrine ; Rats ; Rats, Sprague-Dawley ; Receptors, Adrenergic, alpha-2/metabolism* ; Substance Withdrawal Syndrome/drug therapy* ; Yohimbine/pharmacology*

Spinal alpha-2 adrenoceptors and cholinergic receptors are involved in the regulation of acute nociception and the facilitated processing. The aim of this study was to examine the pharmacological effect of an intrathecal alpha-2 agonist and a cholinesterase inhibitor on the facilitated pain model induced by formalin injection and to determine the nature of drug interaction using an isobolographic analysis. Both intrathecal clonidine and neostigmine dose-dependently suppressed the flinching during phase 1 and phase 2. Intrathecal pretreatment with atropine reversed the antinociceptive effects of clonidine and neostigmine in both phases. Pretreatment with intrathecal yohimbine attenuated the effect of clonidine. The antinociception of clonidine and neostigmine was not reversed by mecamylamine. Isobolographic analysis showed that intrathecal clonidine and neostigmine acted synergistically in both phase 1 and 2. Intrathecal pretreatment with atropine and yohimbine antagonized the effect of the mixture of clonidine and neostigmine in both phases, but no antagonism was observed with mecamylamine pretreatment. These data indicate that spinal clonidine and neostigmine are effective to counteract the facilitated state evoked formalin stimulus, and these two drugs interact in a synergistic fashion. In addition, the analgesic action of intrathecal clonidine is mediated by spinal muscarinic receptors as well as alpha-2 adrenoceptors.
Adrenergic alpha-Agonists/*pharmacology ; Analgesics, Non-Narcotic/*pharmacology ; Animal ; Cholinesterase Inhibitors/*pharmacology ; Clonidine/administration & dosage/*pharmacology ; Dose-Response Relationship, Drug ; Drug Synergism ; Formaldehyde ; Injections, Spinal ; Male ; Neostigmine/administration & dosage/*pharmacology ; Pain/drug therapy ; Rats ; Rats, Sprague-Dawley

It is known that stimulation of the α(2A)-adrenoceptors (α(2A)-ARs) by the selective α(2A)-AR agonist guanfacine produces an important and beneficial influence on prefrontal cortical (PFC) cognitive functions such as spatial working memory and selective attention. However, it is unclear whether stimulation of the α(2A)-ARs has a similar effect on fear conditioning that involves the amygdala and hippocampus. Here, we show that systemically administered guanfacine significantly enhances spatial learning of rats in the Lashley maze: compared with controls, the rats treated with guanfacine required significantly fewer trials and made significantly fewer errors to reach learning criterion. However, guanfacine produced no effect on acquisition of contextual and auditory fear memories. The present study suggests that beneficial effect of α(2A)-AR stimulation is task-dependent: guanfacine improves spatial learning but not fear conditioning.
Adrenergic alpha-2 Receptor Agonists ; pharmacology ; Animals ; Behavior, Animal ; drug effects ; Conditioning (Psychology) ; drug effects ; Fear ; drug effects ; Guanfacine ; pharmacology ; Maze Learning ; drug effects ; Memory ; drug effects ; Rats ; Spatial Behavior ; drug effects

Objective: To explore the protective effects of dexmedetomidine (Dex) against high glucose-induced epithelial-mesenchymal transition in HK-2 cells and relevant mechanisms. Methods: HK-2 cells were exposed to either glucose or glucose+Dex for 6 h. The production of ROS, morphology of HK-2 cells, and cell cycle were detected. Moreover, the expression of AKT, p-AKT, ERK, p-ERK, PI3K, E-Cadherin, Claudin-1, and α-SMA were determined and compared between HK-2 cells exposed to glucose and those exposed to both glucose and Dex with or without PI3K/AKT pathway inhibitor LY294002 and ERK pathway inhibitor U0126. Results: Compared with HK-2 cells exposed to high level of glucose, the HK-2 cells exposed to both high level of glucose and Dex showed: (1) lower level of ROS production; (2) cell morphology was complete; (3) more cells in G1 phase; (4) lower expression of p-AKT, p-ERK and α-SMA, higher expression of E-Cadherin and Claudin-1. PI3K/AKT inhibitor LY294002 and ERK inhibitor U0126 decreased the expression of p-AKT, p-ERK and α-SMA, and increased the expression of E-Cadherin and Claudin-1. Conclusion Dex can attenuate high glucose-induced HK-2 epithelial-mesenchymal transition by inhibiting AKT and ERK.
Adrenergic alpha-2 Receptor Agonists ; pharmacology ; Cell Line ; Dexmedetomidine ; pharmacology ; Epithelial-Mesenchymal Transition ; drug effects ; Glucose ; metabolism ; Humans ; MAP Kinase Signaling System ; drug effects ; Proto-Oncogene Proteins c-akt ; antagonists & inhibitors ; Signal Transduction ; drug effects

We sought to determine the effects of activation of peroxisome proliferator-activated receptor-gamma (PPAR-gamma) on multilocularization of adipocytes in adult white adipose tissue (WAT). Male C57BL/6 normal, db/db, and ob/ob mice were treated with agonists of PPAR-gamma, PPAR-alpha, or beta3-adrenoceptor for 3 weeks. To distinguish multilocular adipocytes from unilocular adipocytes, whole-mounted adipose tissues were co-immunostained for perilipin and collagen IV. PPAR-gamma activation with rosiglitazone or pioglitazone induced a profound change of unilocular adipocytes into smaller, multilocular adipocytes in adult WAT in a time-dependent, dose-dependent, and reversible manner. PPAR-alpha activation with fenofibrate did not affect the number of locules or remodeling. db/db and ob/ob obese mice exhibited less multilocularization in response to PPAR-gamma activation compared to normal mice. Nevertheless, all adipocytes activated by PPAR-gamma contained a single nucleus regardless of locule number. Multilocular adipocytes induced by PPAR-gamma activation contained substantially increased mitochondrial content and enhanced expression of uncoupling protein-1, PPAR-gamma coactivator-1-alpha , and perilipin. Taken together, PPAR-gamma activation induces profound multilocularization and enhanced mitochondrial biogenesis in the adipocytes of adult WAT. These changes may affect the overall function of WAT.
Adipocytes/*cytology/metabolism ; Adipose Tissue, White/*cytology ; Animals ; Cell Nucleus Division ; Hypoglycemic Agents/pharmacology ; Ion Channels/metabolism ; Male ; Mice ; Mice, Inbred C57BL ; Mice, Obese ; Mitochondria/metabolism ; Mitochondrial Proteins/metabolism ; PPAR alpha/agonists/metabolism ; PPAR gamma/*agonists/*metabolism ; Phosphoproteins/metabolism ; Receptors, Adrenergic, beta-3/agonists ; Thiazolidinediones/pharmacology ; Trans-Activators/metabolism