Atsushi Sanbe - Academia.edu (original) (raw)

Papers by Atsushi Sanbe

Current eye research, 2015

To examine whether astaxanthin (AST) prevent the cataract formation induced by glucocorticoid in ... more To examine whether astaxanthin (AST) prevent the cataract formation induced by glucocorticoid in chick embryo. Hydrocortisone hemisuccinate sodium (HC) (0.5 μmol/egg) was administered directly into the air chamber in the egg shell of chick embryo day 15. The eggs were then kept in an incubator at same conditions and administered 100 μL of 50 (HC + AST50 group), 80 (HC + AST80 group), 100 (HC + AST100 group) mg/mL of AST solutions dissolved in dimethyl sulfoxide (DMSO) 3 h after administration of HC. In addition, non-HC treated group (treated with physiological saline without HC and 100 μL of DMSO), HC-alone group (treated with 0.5 μmol of HC and 100 μL of DMSO), and AST100 group (treated with physiological saline without HC and 100 μL of DMSO) were also incorporated. After 48 h of treatment, lenses were removed from embryo and classified into five stages according to developed opacity. The amounts of reduced glutathione in the lenses and the blood glucose levels were measured. The a...

Biological & Pharmaceutical Bulletin, 2015

We previously reported that serotonin (5-hydroxytryptamine; 5-HT) suppresses β-casein expression,... more We previously reported that serotonin (5-hydroxytryptamine; 5-HT) suppresses β-casein expression, a differentiation marker in mammary epithelial cells, via inhibition of the signal transducer and activator of transcription 5 (STAT5) phosphorylation in the human mammary epithelial cell line, MCF-12A. In this study, we investigated the expression pattern of the different 5-HT receptor subtypes in MCF-12A cells, and identified the receptors involved in 5-HT-mediated suppression of β-casein protein expression. β-Casein mRNA expression was inhibited by 30 µM 5-HT in a time-dependent manner. Treatment with 30 µM 5-HT for 72 h decreased β-casein protein levels and STAT5 phosphorylation (pSTAT5). The cells expressed four 5-HT receptors subtypes (5-HTR 1D, 2B, 3A, and 7) at the mRNA and protein level, and their expression was elevated by prolactin (PRL) treatment. Additionally, the mRNA levels of 5-HTR 1D and 5-HTR 7 were significantly higher than the other 5-HT receptors in the cells. Tryptophan hydroxylase 1 mRNA was detectable in the cells in the absence of PRL, and PRL treatment significantly increased its expression. β-Casein and pSTAT5/STAT5 levels in the cells co-treated with 5-HT and a selective 5-HTR 1D inhibitor, BRL15572, were equal to those observed in cells treated with 5-HT alone. However, in the cells co-treated with 5-HT and a selective 5-HTR 7 inhibitor, SB269970, β-casein and pSTAT5/STAT5 levels increased in a SB269970 concentration-dependent manner. In conclusion, we showed that 5-HT regulates β-casein expression via 5-HTR 7 in MCF-12A human mammary epithelial cells.

Cellular Signalling, 2012

Background: The Arf6 activator, cytohesin-2, is involved in neurite growth. Results: Cytohesin-2 ... more Background: The Arf6 activator, cytohesin-2, is involved in neurite growth. Results: Cytohesin-2 binds to CCDC120 and is transported along growing neurites. Conclusion: This interaction is required for Arf6 activation and neurite growth. Significance: The previously unknown functional CCDC120 is a new cytohesin adaptor protein, which regulates neurite growth. The mechanism of neurite growth is complicated, involving continuous cytoskeletal rearrangement and vesicular trafficking. Cytohesin-2 is a guanine nucleotide exchange factor for Arf6, an Arf family molecular switch protein, controlling cell morphological changes such as neuritogenesis. Here, we show that cytohesin-2 binds to a protein with a previously unknown function, CCDC120, which contains three coiled-coil domains, and is transported along neurites in differentiating N1E-115 cells. Transfection of the small interfering RNA (siRNA) specific for CCDC120 into cells inhibits neurite growth and Arf6 activation. When neurites start to extend, vesicles containing CCDC120 and cytohesin-2 are transported in an anterograde manner rather than a retrograde one. As neurites continue extension, anterograde vesicle transport decreases. CCDC120 knockdown inhibits cytohesin-2 localization into vesicles containing CCDC120 and diffuses cytohesin-2 in cytoplasmic regions, illustrating that CCDC120 determines cytohesin-2 localization in growing neurites. Reintroduction of the wild type CCDC120 construct into cells transfected with CCDC120 siRNA reverses blunted neurite growth and Arf6 activity, whereas the cytohesin-2-binding CC1 region-deficient CCDC120 construct does not. Thus, cytohesin-2 is transported along neurites by vesicles containing CCDC120, and it mediates neurite growth. These results suggest a mechanism by which guanine nucleotide exchange factor for Arf6 is transported to mediate neurite growth.

Journal of Molecular and Cellular Cardiology, 1995

Although pharmacological therapy with angiotensin converting enzyme (ACE) inhibitors has proved t... more Although pharmacological therapy with angiotensin converting enzyme (ACE) inhibitors has proved to be effective in patients with heart failure (HF), the experimental basis of this effect has not yet been addressed. In the present study, animals with HF were treated with an oral administration of 10 mg/kg/day captopril, 10 mg/kg/day enalapril and 3 mg/kg/day trandolapril from the 2nd to 12th week after the operation. HF was induced by permanent occlusion of the left coronary artery of the rat at 2 mm from its origin. Treatment of the HF rats with the ACE inhibitors enhanced the decrease in mean arterial blood pressure, attenuated the rise in left ventricular end-diastolic pressure, an indirect marker of preload, and diminished the reduction in cardiac output and stroke volume indices of the HF animal. Treatment also reversed the reduction in ATP, creatine phosphate, creatine and the mitochondrial oxygen consumption rate of the viable left and right ventricles of the HF animal. The improvement of the cardiac output index and high-energy phosphate levels of the HF rat by the ACE inhibitors was associated with the recovery of the mitochondrial oxygen consumption rate. In sham-operated animals, treatment with the ACE inhibitors reduced mean arterial pressure and left ventricular systolic pressure, but not metabolic variables concerning myocardial energy metabolism. The present results provide evidence that ACE inhibitor therapy improves cardiac function and myocardial energy metabolism of experimental animals with chronic heart failure. The mechanism underlying the benefit of long-term treatment with ACE inhibitors is probably attributable to recovery or preservation of the mitochondrial function and reduction in preload.

Naunyn-Schmiedeberg's Archives of Pharmacology, 2009

Maternal smoking during the perinatal period is linked to adverse neonatal outcomes such as low b... more Maternal smoking during the perinatal period is linked to adverse neonatal outcomes such as low birth weight and birth defects. Numerous studies have shown that cigarette smoke or nicotine exposure has a widespread effect on fetal nerve development. However, there exists a lack of understanding of what specific changes occur at the cellular level on persistent exposure to cigarette smoke during the differentiation of embryonic stem cells (ESCs) into neural cells. We previously investigated the effects of cigarette smoke extract (CSE) and its major component, nicotine, on the neural differentiation of mouse embryonic stem cells (mESCs). Differentiation of mESCs into neural progenitor cells (NPCs) or neural crest cells (NCCs) was induced with chemically defined media, and the cells were continuously exposed to CSE or nicotine during neural differentiation and development. Disturbed balance of the pluripotency state was observed in the NPCs, with consequent inhibition of neurite outgrowth and glial fibrillary acidic protein (Gfap) expression. These inhibitions correlated with the altered expression of proteins involved in the Notch-1 signaling pathways. The migration ability of NCCs was significantly decreased by CSE or nicotine exposure, which was associated with reduced protein expression of migration-related proteins. Taken together, we concluded that CSE and nicotine inhibit differentiation of mESCs into NPCs or NCCs, and may disrupt functional development of neural cells. These results imply that cigarette smoking during the perinatal period potentially inhibits neural differentiation and development of ESCs cells, leading to neonatal abnormal brain development and behavioral abnormalities.

Proceedings for Annual Meeting of The Japanese Pharmacological Society, 2022

Proceedings for Annual Meeting of The Japanese Pharmacological Society, 2020

It is known that Bcl-2 associated athanogene (BAG) 3 is strongly expressed in cardiac muscle as w... more It is known that Bcl-2 associated athanogene (BAG) 3 is strongly expressed in cardiac muscle as well as skeletal muscle. BAG3 can directly bind to Bcl-2 as well as heat shock protein (HSP) as a co-chaperone. Recent study showed that myofibrillar degeneration, disruption of Z-disk architecture and apoptotic cell death were observed in BAG3 knockout mouse. Thus, BAG3 may play a protective role in the muscles. We examined BAG3 protein levels in alpha-B crystallin (CryAB) R120G transgenic (TG) mouse, myofibrillar myopathy (MFM) model. A marked increase in BAG3 was observed in MFM hearts. Little is known, however, detail roles of the increased BAG3 in cardiac muscle. In order to understand functional role of increased cardiac BAG3 in MFM hearts, CryAB R120G TG mice were crossbred with TG mice overexpressing BAG3 to generate CryAB R120G/BAG3 double TG mice. Decrease in fractional shortening and induction of cardiac ANP as well as increase in heart weight/body weight ratio were seen in CryAB R120G TG mice. Moreover, deterioration in cardiac function as well as enhanced cardiac hypertrophy were observed in the CryAB R120G/BAG3 double TG mice. Thus, cardiac BAG3 overexpression may be insufficient for prevention of cardiac disease in CryAB R120G TG mice.

Folia Pharmacologica Japonica, 2015

Biological and Pharmaceutical Bulletin, 2014

Serotonin (5-hydroxytryptamine; 5-HT) has an important physiological role in controlling lactatio... more Serotonin (5-hydroxytryptamine; 5-HT) has an important physiological role in controlling lactation, namely, milk volume homeostasis, within mammary glands. The objectives of this study were to evaluate whether exogenous 5-HT can suppress β-casein expression, a differentiation marker, produced in human mammary epithelial cells, and to determine whether 5-HT can attenuate β-casein signaling via the prolactin (PRL) receptor (PRLr) and Janus kinase 2/signal transducer and activator of transcription 5 (STAT5) pathway. PRL treatment increased the mRNA level of β-casein in the MCF-12A human mammary epithelial cell line, and the highest level occurred at days 7 and 14 of culture. In contrast, PRLr expression was not affected significantly by PRL treatment. PRL treatment in MCF-12A cells increased levels of β-casein and phosphorylated STAT5 (pSTAT5) proteins in a concentration-dependent manner, with a slight increase of STAT5 protein. β-Casein expression was inhibited by 0.1 mM 5-HT in a time-dependent manner. Additionally, treatment with 0.1 mM 5-HT for 72 h decreased protein levels of β-casein and pSTAT5, with a slight decrease in STAT5 levels. These results suggest that exogenous 5-HT can inhibit STAT5 phosphorylation, resulting in a decrease in β-Casein expression. In conclusion, we showed that exogenous 5-HT decreased β-casein expression in MCF-12A human mammary epithelial cells, and that 5-HT was responsible for inhibiting phosphorylation of STAT5, resulting in a decline in lactational function.

PLoS ONE, 2013

Neural cell differentiation during development is controlled by multiple signaling pathways, in w... more Neural cell differentiation during development is controlled by multiple signaling pathways, in which protein phosphorylation and dephosphorylation play an important role. In this study, we examined the role of pyrophosphatase1 (PPA1) in neuronal differentiation using the loss and gain of function analysis. Neuronal differentiation induced by external factors was studied using a mouse neuroblastoma cell line (N1E115). The neuronal like differentiation in N1E115 cells was determined by morphological analysis based on neurite growth length. In order to analyze the loss of the PPA1 function in N1E115, si-RNA specifically targeting PPA1 was generated. To study the effect of PPA1 overexpression, an adenoviral gene vector containing the PPA1 gene was utilized to infect N1E115 cells. To address the need for pyrophosphatase activity in PPA1, D117A PPA1, which has inactive pyrophosphatase, was overexpressed in N1E115 cells. We used valproic acid (VPA) as a neuronal differentiator to examine the effect of PPA1 in actively differentiated N1E115 cells. Si-PPA1 treatment reduced the PPA1 protein level and led to enhanced neurite growth in N1E115 cells. In contrast, PPA1 overexpression suppressed neurite growth in N1E115 cells treated with VPA, whereas this effect was abolished in D117A PPA1. PPA1 knockdown enhanced the JNK phosphorylation level, and PPA1 overexpression suppressed it in N1E115 cells. It seems that recombinant PPA1 can dephosphorylate JNK while no alteration of JNK phosphorylation level was seen after treatment with recombinant PPA1 D117A. Enhanced neurite growth by PPA1 knockdown was also observed in rat cortical neurons. Thus, PPA1 may play a role in neuronal differentiation via JNK dephosphorylation.

The Journal of Physiology, 2007

We have reported that [Arg 8 ]-vasopressin-stimulated insulin release is blunted in islet cells i... more We have reported that [Arg 8 ]-vasopressin-stimulated insulin release is blunted in islet cells isolated from V1b receptor-deficient (V1bR −/−) mice. In this study, we used V1bR −/− mice to examine the physiological role of the V1b receptor in regulating blood glucose levels in vivo, and we found that the fasting plasma glucose, insulin and glucagon levels were lower in V1bR −/− mice than in wild-type (V1bR +/+) mice. Next, we evaluated glucose tolerance by performing an intraperitoneal glucose tolerance test (GTT). The plasma glucose and insulin levels during the GTT were lower in V1bR −/− mice than in V1bR +/+ mice. An insulin tolerance test (ITT) revealed that, after insulin administration, plasma glucose levels were lower in V1bR −/− mice than in V1bR +/+ mice. In addition, a hyperinsulinaemic-euglycaemic clamp study showed that the glucose infusion rate was increased in V1bR −/− mice, indicating that insulin sensitivity was enhanced at the in vivo level in V1bR −/− mice. Furthermore, we found that the V1b receptor was expressed in white adipose tissue and that insulin-stimulated phosphorylation of Akt as an important signaling molecule was increased in adipocytes isolated from V1bR −/− mice. Thus, the blockade of the V1b receptor could result, at least in part, in enhanced insulin sensitivity by altering insulin signalling in adipocytes.

PLoS ONE, 2011

Background: Transgenic (TG) mice with overexpression of an arg120gly (R120G) missense mutation in... more Background: Transgenic (TG) mice with overexpression of an arg120gly (R120G) missense mutation in HSPB5 display desmin-related cardiomyopathy, which is characterized by formation of aggresomes. It is also known that progressive mitochondrial abnormalities and apoptotic cell death occur in the hearts of R120G TG mice. The role of mitochondrial dysfunction and apoptosis in disease progression, however, remains uncertain. Methods and Results: Mitochondrial abnormalities and apoptotic cell death induced by overexpression of HSPB5 R120G were analyzed in neonatal rat cardiomyocytes. Overexpression of mutant HSPB5 led to development of aggresomes with a concomitant reduction in cell viability in the myocytes. Overexpression of mutant HSPB5 induced a reduction in the cytochrome c level in the mitochondrial fraction and a corresponding increase in the cytoplasmic fraction in the myocytes. Down-regulation of BCL2 and up-regulation of BAX were detected in the myocytes expressing the mutant HSPB5. Concomitant with mitochondrial abnormality, the activation of caspase-3 and increased apoptotic cell death was observed. Cell viability was dose-dependently recovered in myocytes overexpressing HSPB5 R120G by treatment with nicorandil a mitochondrial ATP-sensitive potassium channel opener. Nicorandil treatment also inhibited the increase in BAX, the decrease in BCL2, activation of caspase-3 and apoptotic cell death by mutant HSPB5. To confirm the results of the in-vitro study, we analyzed the effect of nicorandil in HSPB5 R120G TG mice. Nicorandil treatment appeared to reduce mitochondrial impairment and apoptotic cell death and prolonged survival in HSPB5 R120G TG mice. Conclusions: Nicorandil may prolong survival in HSPB5 R120G TG mice by protecting against mitochondrial impairments.

Journal of Molecular and Cellular Cardiology, 2006

Although it has been suggested that in cardiac muscle the phosphorylation level of myosin regulat... more Although it has been suggested that in cardiac muscle the phosphorylation level of myosin regulatory light chain (RLC) correlates with frequency of stimulation, its significance in the modulation of the force-frequency and pressure-frequency relationships remains unclear. We examined the role of RLC phosphorylation on the force-frequency relation (papillary muscles), the pressure-frequency relation (Langendorff perfused hearts) and shortening-frequency relation (isolated cardiac myocytes) in nontransgenic (NTG) and transgenic mouse hearts expressing a nonphosphorylatable RLC protein (RLC(P−)). At 22°C, NTG and RLC(P−) muscles showed a negative force-frequency relation. At 32°C, at frequencies above 1 Hz, both groups showed a flat force-frequency relation. There was a small increase in RLC phosphorylation in NTG muscles when the frequency of stimulation was increased from 0.2 Hz to 4.0 Hz. However, the level of RLC phosphorylation in these isolated muscles was significantly lower compared to samples taken from NTG intact hearts. In perfused hearts, there was no difference in the slope of pressurefrequency relationship between groups, but the RLC(P−) group consistently developed a reduced systolic pressure and demonstrated a decreased contractility. There was no difference in the level of RLC phosphorylation in hearts paced at 300 and 600 bpm. In RLC(P−) hearts, the level of TnI phosphorylation was reduced compared to NTG. There was no change in the expression of PLB between groups, but expression of SERCA2 was increased in hearts from RLC(P−) compared to NTG. In isolated cardiac myocytes, there was no change in shortening-frequency relationship between groups. Moreover, there was no change in Ca 2+ transient parameters in cells from NTG and RLC(P−) hearts. Our data demonstrate that in cardiac muscle RLC phosphorylation is not an essential determinant of force-and pressure-frequency relations but the absence of RLC phosphorylation decreases contractility in force/pressure developing preparations.

Journal of Endocrinology, 2007

Arg 8 ]-vasopressin (AVP) and oxytocin (OT) are neurohypophysial hormones which exert various act... more Arg 8 ]-vasopressin (AVP) and oxytocin (OT) are neurohypophysial hormones which exert various actions, including the control of blood glucose, in some peripheral tissues. To investigate the type of receptors involved in AVP-and OT-induced glucagon secretion, we investigated the effect of these peptides on glucagon secretion in islets of wild-type (V1bRC/C) and vasopressin V1b receptor knockout (V1bRK/K) mice. AVP-induced glucagon secretion was significantly inhibited by the selective V1b receptor antagonist, SSR149415 (30%), and OT-induced glucagon secretion by the specific OT receptor antagonist, dðCH 2 Þ 5 ½TyrðMeÞ 2 ; Thr 4 ; Tyr-NH 9 2 OVT (CL-14-26) (45%), in islets of V1bRC/Cmice. AVP-and OT-induced glucagon secretions were not by the antagonist of each, but co-incubation with both 10 K6 M SSR149415 and 10 K6 M CL-14-26 further inhibited AVP-and OT-induced glucagon secretions in islets of V1bRC/C mice (57 and 69% of the stimulation values respectively). In addition, both AVP and OT stimulated glucagon secretion with the same efficacy in V1bRK/K mice as in V1bRC/C mice. AVP-and OT-induced glucagon secretion in V1bRK/K mice was significantly inhibited by CL-14-26. These results demonstrate that V1b receptors can mediate OT-induced glucagon secretion and OTreceptors can mediate AVP-induced glucagon secretion in islets from V1bRC/C mice in the presence of a heterologous antagonist, while AVP and OT can stimulate glucagon secretion through the OT receptors in V1bRK/K mice, suggesting that the other receptor can compensate when one receptor is absent.

Journal of Biological Chemistry, 2013

Background: Dock4, a guanine nucleotide exchange factor for Rac1, is associated with neuropsychia... more Background: Dock4, a guanine nucleotide exchange factor for Rac1, is associated with neuropsychiatric diseases. Results: Dock4 regulates neurite differentiation in neuroblastoma cells and hippocampal neurons. Conclusion: Dock4 is an important regulator during neural differentiation. Significance: This study contributes to a better understanding of the molecular and cellular events during neural differentiation and may provide new insights into the molecular pathophysiology of neuropsychiatric diseases. Precise regulation of neurite growth and differentiation determines accurate formation of synaptic connections, whose disruptions are frequently associated with neurological disorders. Dedicator of cytokinesis 4 (Dock4), an atypical guanine nucleotide exchange factor for Rac1, is found to be associated with neuropsychiatric diseases, including autism and schizophrenia. Nonetheless, the neuronal function of Dock4 is only beginning to be understood. Using mouse neuroblastoma (Neuro-2a) cells as a model, this study identifies that Dock4 is critical for neurite differentiation and extension. This regulation is through activation of Rac1 and modulation of the dynamics of actin-enriched protrusions on the neurites. In cultured hippocampal neurons, Dock4 regulates the establishment of the axon-dendrite polarity and the arborization of dendrites, two critical processes during neural differentiation. Importantly, a microdeletion Dock4 mutant linked to autism and dyslexia that lacks the GEF domain leads to defective neurite outgrowth and neuronal polarization. Further analysis reveals that the SH3 domain-mediated interaction of Dock4 is required for its activity toward neurite differentiation, whereas its proline-rich C terminus is not essential for this regulation. Together, our findings reveal an important role of Dock4 for neurite differentiation during early neuronal development.

Journal of Biological Chemistry, 2010

The formation of primitive adipose tissue is the initial process in adipose tissue development fo... more The formation of primitive adipose tissue is the initial process in adipose tissue development followed by the migration of preadipocytes into adipocyte clusters. Comparatively little is known about the molecular mechanism controlling preadipocyte migration. Here, we show that cytohesin-2, the guaninenucleotide exchange factor for the Arf family GTP-binding proteins, regulates migration of mouse preadipocyte 3T3-L1 cells through Arf6. SecinH3, a specific inhibitor of the cytohesin family, markedly inhibits migration of 3T3-L1 cells. 3T3-L1 cells express cytohesin-2 and cytohesin-3, and knockdown of cytohesin-2 with its small interfering RNA effectively decreases cell migration. Cytohesin-2 preferentially acts upstream of Arf6 in this signaling pathway. Furthermore, we find that the focal adhesion protein paxillin forms a complex with cytohesin-2. Paxillin colocalizes with cytohesin-2 at the leading edges of migrating cells. This interaction is mediated by the LIM2 domain of paxillin and the isolated polybasic region of cytohesin-2. Importantly, migration is inhibited by expression of the constructs containing these regions. These results suggest that cytohesin-2, through a previously unexplored complex formation with paxillin, regulates preadipocyte migration and that paxillin plays a previously unknown role as a scaffold protein of Arf guanine-nucleotide exchange factor.

Journal of Biological Chemistry, 2001

The functional significance of the actin binding region at the amino terminus of the cardiac esse... more The functional significance of the actin binding region at the amino terminus of the cardiac essential myosin light chain (ELC) remains obscure. Previous experiments carried out in vitro indicated that modulation of residues 5-14 could induce an inotropic effect, increasing maximal ATPase activity at submaximal Ca 2؉ concentrations (Rarick, H.

Journal of Biological Chemistry, 2006

An R120G missense mutation in ␣-B-crystallin (CryAB), a small heat-shock protein (HSP), causes a ... more An R120G missense mutation in ␣-B-crystallin (CryAB), a small heat-shock protein (HSP), causes a desmin-related cardiomyopathy (DRM) that is characterized by the formation of aggregates containing CryAB and desmin. The mutant CryAB protein leads to the formation of inclusion bodies, which contain amyloid oligomer intermediates (amyloid oligomer) in the cardiomyocytes. To further address the underlying mechanism(s) of amyloid oligomer formation in DRM linked to the CryAB R120G, a recombinant CryAB R120G protein was generated. The purified CryAB R120G protein can form a toxic amyloid oligomer, whereas little immunoreactivity was observed in the wild-type CryAB protein. A native PAGE showed that the oligomerized form was present in the CryAB R120G protein, whereas only a high molecular mass was detected in the wildtype CryAB. The oligomerized CryAB R120G of around 240-480 kDa showed strong positive immunoreactivity against an anti-oligomer antibody. The CryAB R120G amyloid oligomer was unstable and easily lost its conformation by ␤-mercaptoethanol and SDS. Recombinant HSP25 or HSP22 proteins can directly interrupt oligomer formation by the CryAB R120G protein, whereas the amyloid oligomer is still present in the mixture of the wild-type CryAB and CryAB R120G proteins. This interruption by HSP25 and HSP22 was confirmed in a cardiomyocyte-based study using an adenoviral transfection system. Blockade of amyloid oligomer formation by HSP25 and HSP22 recovered the ubiquitin proteosomal activity and cellular viability. Blockade of oligomer formation by small HSP may be a new therapeutic strategy for treating DRM as well as other types of amyloid-based degenerative diseases.

Investigative Opthalmology & Visual Science, 2011

PURPOSE. To identify the ␣ 1-adrenoceptor (␣ 1-AR) subtypes mediating vascular adrenergic respons... more PURPOSE. To identify the ␣ 1-adrenoceptor (␣ 1-AR) subtypes mediating vascular adrenergic responses in murine ophthalmic arteries. METHODS. Expression of mRNA was quantified for individual ␣ 1-AR subtypes in murine ophthalmic arteries using real-time PCR. To assess the functional relevance of ␣ 1-ARs for mediating vascular responses, ophthalmic arteries from mice deficient in one of the three ␣ 1-AR subtypes (␣ 1A-AR Ϫ/Ϫ , ␣ 1B-AR Ϫ/Ϫ , and ␣ 1D-AR Ϫ/Ϫ , respectively) and wild-type controls were isolated, cannulated with micropipettes, and pressurized. Changes in luminal artery diameter in response to the ␣ 1-AR agonist phenylephrine, the sympathetic transmitter noradrenaline, and to the nonadrenergic vasoconstrictor arginine vasopressin (AVP) were measured by video microscopy. RESULTS. Using real-time PCR, mRNA for all three ␣ 1-AR subtypes was detected in ophthalmic arteries from wild-type mice. In functional studies, phenylephrine and noradrenaline produced dose-dependent constriction of ophthalmic arteries that was similar in wild-type, ␣ 1B-AR Ϫ/Ϫ , and ␣ 1D-AR Ϫ/Ϫ mice. Strikingly, responses to phenylephrine and noradrenaline were almost completely abolished in ␣ 1A-AR Ϫ/Ϫ mice. In contrast, the nonadrenergic agonist AVP produced dose-dependent vasoconstrictor responses that did not differ between any of the mouse genotypes tested. CONCLUSIONS. These findings provide evidence that the ␣ 1A-AR subtype mediates adrenergic vasoconstriction in murine ophthalmic arteries.