摘要
核苷酸类物质对胃肠道动力和分泌功能具有重要的调节作用,其中ATP主要通过P2受体介导胃肠道平滑肌的收缩与舒张反应。有研究发现ATP诱发大鼠近端胃环行肌先舒张后收缩的双相反应。而本室研究曾报道,ATP仅使胃体环行肌产生收缩反应。我们推测,大鼠胃体环行肌的药理学特征可能不同与胃底环行肌,是分析P2受体介导胃平滑肌收缩反应的理想实验标本。P2受体包括P2X1-7和P2Y1-14多种亚型,功能复杂。目前,有关核苷酸类物质诱发大鼠胃体环行肌收缩反应的P2Y受体亚型分析、大鼠胃体平滑肌P2Y受体亚型mRNA表达的绝对定量分析及P2Y1与P2Y2受体在大鼠胃体平滑肌的表达,尚未见文献报道。本实验研究了P2受体介导大鼠胃体环行肌收缩反应的药理学特点,并应用实时荧光定量PCR技术绝对定量分析大鼠胃体平滑肌P2Y受体亚型mRNA的表达和免疫组织化学技术分析大鼠胃体平滑肌P2Y1与P2Y2受体的表达。
第一部分核苷及核苷酸类物质介导大鼠胃体环行肌收缩反应的药理学特点
制备大鼠离体胃体和胃底环行肌标本。利用受体药理学技术观察大鼠胃体和胃底环行肌不同的药理学特征,并进一步分析核苷及核苷酸类物质诱发胃体环行肌收缩反应的作用特点。
1 CCh、5-HT、His和KCl对大鼠胃体环行肌和胃底环行肌的作用
在胃体环行肌KCl所致收缩反应与胃底环行肌无显著性差别;但是,CCh收缩胃体环行肌的EC50值(0.45±0.15μmol?L-1)显著高于胃底环行肌(0.20±0.09μmol?L-1,P<0.01)。5-HT和His收缩两种标本的EC50值无显著差异(P>0.05);但是,在胃体环行肌5-HT和His产生收缩反应的Emax值(0.81±0.26和0.88±0.27 g)显著小于胃底环行肌(2.67±0.61和1.90±0.68 g,P<0.01)。
2 ATP对预收缩大鼠胃体环行肌和胃底环行肌的作用
在预收缩胃体环行肌,ATP(0.1-3000μmol?L-1)诱发浓度依赖性收缩反应,未见舒张反应;在预收缩胃底环行肌标本,同浓度ATP诱发先舒张后收缩的双相反应,并呈浓度依赖性。
3核苷及核苷酸对大鼠胃体环行肌的作用
ATP、UTP、ADP、2-MeSATP和α,β-MeATP浓度依赖性诱发大鼠胃体环行肌收缩反应,2-MeSATP的EC50值为7.2±5.2 nmol?L-1比ACh(3.47±1.20μmol?L-1)低500倍;各药物产生收缩反应的效价序列为:2-MeSATP>>ADP>ATP=UTP>α,β-MeATP>>腺苷。
4酚妥拉明、普萘洛尔、阿托品及河豚毒素对ATP和UTP诱发大鼠胃体环行肌收缩反应的影响
酚妥拉明(1μmol?L-1)、普萘洛尔(1μmol?L-1)、阿托品(1μmol?L-1)及河豚毒素(0.1μmol?L-1)不影响ATP和UTP诱发的胃体环行肌收缩反应。
上述结果表明,大鼠胃体环行肌的药理学特征明显不同于胃底环行肌;核苷及核苷酸类物质诱发大鼠收缩反应的效价序列为2-MeSATP>> ADP> ATP= UTP>α,β-MeATP>>腺苷,提示通过P2Y受体介导胃体环行肌收缩反应;核苷酸类物质是调节胃体环行肌收缩功能的重要介质。
第二部分P2Y受体介导大鼠胃体环行肌收缩反应的受体药理学分析
本研究以2-MeSATP和UTP分别脱敏相应的P2Y受体,观察核苷酸类物质诱发大鼠胃体环行肌收缩反应的变化;同时分析了P2受体阻断剂苏拉明和PPADS对核苷酸类物质诱发大鼠胃体环行肌收缩反应的影响。1 UTP和2-MeSATP脱敏P2Y受体对药物诱发大鼠胃体环行肌收缩反应的影响
ATP(30μmol?L-1)、UTP(3μmol?L-1)、ADP(3μmol?L-1)、2-MeSATP(0.03μmol?L-1)、α,β-MeATP(3μmol?L-1)、ACh(3μmol?L-1)、His(30μmol?L-1)和5-HT(3μmol?L-1)诱发大鼠胃体环行肌的收缩反应在给予溶媒前后均无显著性差异(P>0.05)。UTP(9μmol?L-1)脱敏时,ATP、ADP、2-MeSATP和5-HT诱发的大鼠胃体环行肌收缩反应显著增高(P<0.05),其中ATP组增加了41.1%,ADP组增加了46.7%,2-MeSATP组增加了38.6%,5-HT组增加61.3%;而α,β-MeATP、ACh和His诱发的收缩反应无显著性变化(P>0.05)。2-MeSATP(0.09μmol?L-1)脱敏时,ATP和ADP诱发的大鼠胃体环行肌收缩反应明显降低(P<0.01),其中ATP组降低了86.7%,ADP组的收缩反应完全抑制;UTP、α,β-MeATP、ACh、His和5-HT诱发的收缩反应无显著性变化(P>0.05)。
2苏拉明和PPADS对ATP、UTP和2-MeSATP诱发大鼠胃体环行肌收缩反应的影响
苏拉明(100μmol?L-1)显著抑制ATP(30μmol?L-1)、UTP(3μmol?L-1)和2-MeSATP(0.03μmol?L-1)诱发的大鼠胃体环行肌收缩反应(P<0.01);其中ATP组降低了79.0%,UTP组降低了93.3%,2-MeSATP组降低了87.1%。PPADS(30μmol?L-1)使UTP(3μmol?L-1)诱发大鼠胃体环行肌的收缩反应降低61.6%(P<0.01),而对ATP(30μmol?L-1)和2-MeSATP(0.03μmol?L-1)诱发的收缩反应无明显变化(P>0.05)。
上述结果表明,大鼠胃体环行肌存在多种功能性P2Y受体亚型介导收缩反应;2-MeSATP、ATP和ADP可能激动环行肌细胞的同一P2Y受体亚型产生收缩反应,该收缩反应可被P2Y2和P2Y4受体脱敏所增强。P2Y2和P2Y4受体脱敏对5-HT诱发收缩反应的增强更为明显。
第三部分大鼠胃体平滑肌P2Y受体mRNA表达的定量研究
本研究应用RT-PCR技术分析了大鼠胃体平滑肌P2X受体亚型(P2X1-7)和P2Y受体亚型(P2Y1, 2, 4, 6, 11, 12, 13, 14)mRNA的表达;并进一步应用实时荧光定量PCR技术,绝对定量分析P2Y1, 2, 4, 6, 12, 13, 14受体mRNA在大鼠胃体组织的表达水平。
1 RT-PCR法检测P2X和P2Y受体亚型mRNA在大鼠胃体平滑肌的表达
扩增产物使用2%琼脂糖凝胶电泳分析。结果显示阳性对照β-actin mRNA扩增产物显示与预期相同的特异性条带;P2X1, 2, 4, 5, 7和P2Y1, 2, 4, 6, 12, 13, 14受体mRNA扩增产物在各自相应泳道出现与扩增目的片段大小相同的特异性条带;P2X3, 6和P2Y11受体mRNA扩增产物未出现扩增目的条带。
2实时荧光定量PCR法绝对定量分析大鼠胃体平滑肌P2Y受体mRNA的表达
SYBR-green实时定量PCR检测β-actin mRNA在大鼠胃体平滑肌的表达量为9,809,756.9拷贝数/10ng总RNA,显著高于P2Y受体各亚型mRNA在大鼠胃体平滑肌的拷贝数(P<0.01)。各P2Y受体亚型mRNA在大鼠胃体环行肌表达强度顺序为P2Y2> P2Y1>> P2Y12> P2Y6= P2Y14> P2Y13> P2Y4> P2Y11=0。P2Y2受体和P2Y1受体mRNA在大鼠胃体平滑肌的表达量分别为75,997.5±39,709.3和21,611.5±8,294.7(拷贝数/10 ng总RNA),显著高于其他P2Y受体(P<0.01)。
研究结果表明,除P2X3、P2X6和P2Y11受体亚型外,其他P2X和P2Y受体亚型的mRNA均表达于大鼠胃体平滑肌。大鼠胃体平滑肌P2Y受体亚型mRNA的绝对定量PCR分析结果表明,P2Y2受体和P2Y1受体mRNA是优势表达的P2Y受体,各受体亚型的表达强度顺序为P2Y2> P2Y1 >> P2Y12 > P2Y6 = P2Y14 > P2Y13 > P2Y4。
第四部分P2Y1受体和P2Y2受体在大鼠胃体平滑肌细胞的表达
本研究应用免疫组织化学技术分析大鼠胃体平滑肌细胞(SMCs)P2Y1和P2Y2受体的表达,并对P2Y1和P2Y2受体在大鼠胃底与胃体的分布进行了比较。
阴性对照组中,大鼠胃体和胃底全层标本无特异性着色。在大鼠胃体和胃底,粘膜固有层、粘膜肌、纵行肌、环行肌和肌间神经丛可检测到P2Y1和P2Y2受体免疫反应阳性产物。腺体细胞和神经元的胞浆和平滑肌细胞的细胞核出现强阳性着色。
大鼠胃体环行肌SMCs的P2Y1受体和P2Y2受体的阳性细胞率分别为33.08%±9.93%和45.17%±6.11%,显著高于纵行肌和粘膜肌SMCs的阳性细胞率(P<0.01);大鼠胃底纵行肌、环行肌和粘膜肌SMCs P2Y1受体和P2Y2受体的表达与胃体相似。在大鼠胃体和胃底的纵行肌和环行肌层,P2Y2受体的阳性细胞率显著高于P2Y1受体(P<0.05);大鼠胃体粘膜肌SMCs的表达与此类似。大鼠胃体纵行肌、环行肌和粘膜肌SMCs P2Y2受体的阳性细胞率显著低于胃底同类SMCs的阳性细胞率(P<0.05);胃体纵行肌和粘膜肌SMCs的P2Y1受体阳性细胞率显著低于胃底同类SMCs的阳性细胞率(P<0.05),而在环行肌SMCs间无显著性差别(P>0.05)。
研究结果表明,在大鼠胃体和胃底组织,P2Y1受体和P2Y2受体表达于环行肌、纵行肌和粘膜肌SMCs的细胞核;两种受体在环行肌的表达显著高于纵行肌和粘膜肌。P2Y1受体和P2Y2受体在胃体纵行肌和粘膜肌的表达显著低于胃底。此外,在大鼠胃体、胃底环行肌和纵行肌SMCs,P2Y2受体的表达显著高于P2Y1受体;胃体粘膜肌SMCs的P2Y2表达亦显著高于P2Y1。
结论
核苷酸类物质使大鼠胃体环行肌产生浓度依赖性收缩反应,是调节胃体环行肌收缩活动的重要介质;大鼠胃体环行肌的药理学特征明显不同于胃底环行肌。
大鼠胃体环行肌存在多种功能性P2Y受体亚型介导收缩反应。2-MeSATP、ATP和ADP可能作用于环行肌细胞的同一P2Y受体亚型产生收缩反应,该收缩反应可被P2Y2和P2Y4受体脱敏所增强。
P2Y受体家族中的P2Y1受体和P2Y2受体mRNA在大鼠胃体平滑肌组织呈优势表达,各受体亚型mRNA的表达强度顺序为P2Y2 > P2Y1>> P2Y12 > P2Y6 = P2Y14 > P2Y13 > P2Y4。
大鼠胃体和胃底组织中P2Y1和P2Y2受体在环行肌层SMCs的表达显著高于纵行肌层,两种受体在胃体纵行肌层SMCs的表达显著低于胃底。在大鼠胃体、胃底的环行肌层和纵行肌层SMCs,P2Y2受体的表达显著高于P2Y1受体。
Nucleotides play an important role in the modulation of both secretory and motor functions in the gastrointestinal tract. Among them, ATP acts mainly via P2 receptors to produce relaxation and contraction of smooth muscle in the gastrointestinal tract. It was reported that ATP induced a biphasic response (relaxation followed by a contraction) in the circular muscle of the rat proximal stomach. In our study, however, ATP produced only contractile response in the circular muscle of the rat gastric body. We therefore propose that there is an obvious difference in pharmacological characteristics of the circular smooth muscle between rat gastric body and gastric fundus, and this circular muscle strips might be a useful preparation to analyze P2 receptor-mediated contraction of the stomach. P2 receptors include P2X1-7 and P2Y1-14 receptor subtypes mediating complicated multiple functions. Up to the present time there is no evidence to show the involvement of P2Y receptor subtypes in circular muscle contraction, the absolute quantitative analysis of mRNA expression of P2Y receptor subtypes, and P2Y1 and P2Y2 receptor expression in the rat gastric body smooth muscle layer. In the present study, we investigated the pharmacological characteristics of P2 receptor-mediated contraction of the circular smooth muscle between rat gastric body and gastric fundus, mRNA expression of P2Y receptor subtypes by absolute quantitative real-time PCR analysis, and the immunohistochemical staining of P2Y1 and P2Y2 receptor in the rat gastric body.
PartⅠPharmacological characteristics of contractile responses to nucleosides and nucleotides in circular smooth muscle of the rat gastric body
Isolated circular muscle strips of the rat gastric body and gastric fundus were prepared and their contractile responses to nucleosides and nucleotides were investigated. The difference in pharmacological characteristics of the isolated circular muscle strips between rat gastric body and gastric fundus was analyzed.
1 Responses to CCh, 5-HT, His and KCl in circular muscle strips of the rat gastric body and gastric fundus
There was no significant difference in the responses to KCl of the circular muscle strips between gastric body and gastric fundus, and no difference in EC50 values of 5-HT and His to produce contractile responses between the two kinds of preparations (P>0.05). However, Emax values of contractile responses to 5-HT and His (0.81±0.26 and 0.88±0.27 g) in gastric body were significantly smaller than those in gastric fundus (2.67±0.61 and 1.90±0.68 g, P<0.01). EC50 value of CCh to produce contractile response (0.45±0.15μmol?L-1) in gastric body was significantly higher than that in gastric fundus (0.20±0.09μmol?L-1, P<0.01).
2 Responses to ATP in circular muscle strips of the rat gastric body and gastric fundus precontracted with ACh
In precontracted circular muscle strips of the gastric body, ATP (0.1-3000μmol?L-1) produced only a contractile response in a concentration dependent manner, but the same concentration of ATP induced a biphasic response (relaxation followed by a contraction) in precontracted circular muscle strips of the gastric fundus.
3 Responses to nucleosides and nucleotides in circular muscle strips of the rat gastric body
ATP, UTP, ADP, 2-MeSATP andα,β-MeATP produced contractile responses in circular muscle strips of the rat gastric body in a concentration dependent manner, respectively. The EC50 value of 2-MeSATP (7.2±5.2 nmol?L-1) was about 500 times lower than that of ACh (3.47±1.20μmol?L-1). The rank order of potency was 2-MeSATP>> ADP> ATP= UTP>α,β-MeATP>>adenosine.
4 Effects of phentolamine, propranolol, atropine and tetrodotoxin on the contractile responses to ATP and UTP
The contractile responses to ATP and UTP were not significantly affected by phentolamine (1μmol?L-1), propranolol (1μmol?L-1), atropine (1μmol?L-1) and tetrodotoxin (0.1μmol?L-1).
These results indicate that there is a significant difference in pharmacological characteristics of circular muscle strips between the rat gastric body and gastric fundus. The rank order of potency for the contraction was 2-MeSATP>> ADP> ATP= UTP>α,β-MeATP>> adenosine, indicating that nucleotides produce contractile responses mainly via P2Y receptors, and nucleotides are important mediators responsible for the contraction in circular smooth muscle of the rat gastric body.
PartⅡPharmacological analysis of P2Y receptor subtypes involved in the contraction in circular smooth muscle of the rat gastric body
Isolated circular muscle strips of the rat gastric body were prepared and their contractile responses to nucleotides were investigated. The contractile responses induced by nucleotides were analyzed in the preparations desensitized with 2-MeSATP or UTP, and analyzed with antagonists of suramin and PPADS.
1 Changes in contractile response to P2 receptor agonists in circular muscle strips of the rat gastric body desensitized with 2-MeSATP or UTP
Solvent did not affect the contractile responses to ATP (30μmol?L-1), UTP (3μmol?L-1), ADP (3μmol?L-1), 2-MeSATP (0.03μmol?L-1),α,β-MeATP (3μmol?L-1), ACh (3μmol?L-1), His (30μmol?L-1) and 5-HT (3μmol?L-1) in circular muscle strips of the rat gastric body (P>0.05). The contractile responses to ATP, ADP, 2-MeSATP and 5-HT were significantly increased by a desensitization with UTP (9μmol?L-1, P<0.05), but those toα,β-MeATP, ACh and His were not affected significantly (P>0.05). ATP, ADP, 2-MeSATP and 5-HT related contractions were increased by 41.1%, 46.7%, 38.6% and 61.3% in the preparations desensitized with UTP, respectively. The contractile responses to ATP and ADP were significantly decreased by a desensitization with 2-MeSATP (0.09μmol?L-1, P<0.01), but those to UTP,α,β-MeATP, ACh, His and 5-HT were not affected significantly (P>0.05). ATP and ADP related contractions were decreased by 86.7% and 100% in the preparations desensitized with 2-MeSATP, respectively.
2 Effects of suramin and PPADS on the contractile response to ATP, 2-MeSATP and UTP in circular muscle strips of the rat gastric body
Suramin (100μmol?L-1) significantly inhibited the contractile responses to ATP (30μmol?L-1), UTP (3μmol?L-1) and 2-MeSATP (0.03μmol?L-1) by 79.0%, 93.3% and 87.1%, respectively (P<0.01). PPADS (30μmol?L-1) significantly inhibited the contractile response to UTP (3μmol?L-1) by 61.6% (P<0.01), but did not affect the contractile responses to ATP (30μmol?L-1) and 2-MeSATP (0.03μmol?L-1) significantly (P>0.05).
These results indicate that there are several P2Y receptor subtypes involved in the contractile responses induced by nucleotides in circular muscle of rat gastric body. 2-MeSATP, ATP and ADP might act on a same P2Y receptor subtype regulating the contraction, and which is potentiated by a desensitization of P2Y2 and P2Y4 receptors. Desensitization of P2Y2 and P2Y4 receptors also potentiates the contraction response to 5-HT.
PartⅢQuantitative analysis of mRNA expression of P2Y receptor subtypes in the rat gastric body muscle layer
The mRNA expression of P2X1-7 and P2Y1, 2, 4, 6, 11, 12, 13, 14 receptors was detected by reverse transcription-polymerase chain reaction (RT-PCR) analysis, and the mRNA expression of P2Y1, 2, 4, 6, 12, 13, 14 receptors was further analyzed by absolute quantitative RT-PCR utilizing SYBR-green fluorescence in the rat gastric body muscle layer.
1 The mRNA expression of P2X and P2Y receptor subtypes detected by RT-PCR analysis in the rat gastric body muscle layer
The PCR products were analyzed by electrophoresis on a 2% agarose gel. We detected a single prominent band of the expected size ofβ-actin mRNA after PCR amplification of RNA preparation. Specific bands of the expected sizes of P2X1, 2, 4, 5, 7 and P2Y1, 2, 4, 6, 12, 13, 14 receptor transcripts were detected by agarose gel electrophoresis, but we were not able to confirm the specific bands of the expected sizes of P2X3, 6 and P2Y11 receptor transcripts.
2 The mRNA expressions of P2Y receptor subtypes detected by absolute quantitative real-time PCR analysis in the rat gastric body muscle layer
Theβ-actin mRNA copy number detected by absolute quantitative real-time PCR utilizing SYBR-green fluorescence were 9,809,756.9 copies per 10 ng of total RNA in the rat gastric body muscle layer, which was significantly higher than the mRNA copy number of each P2Y receptor subtype (P<0.01). The relative rank order of P2Y receptor subtype mRNA expression was P2Y2 > P2Y1 >> P2Y12 > P2Y6 = P2Y14 > P2Y13 > P2Y4 > P2Y11 = 0. Expression level of P2Y1 or P2Y2 receptor mRNA was significant higher than that of the other P2Y receptor subtype mRNA in the rat gastric body muscle layer (P<0.01), and P2Y1 and P2Y2 receptor mRNA copy number detected by absolute quantitative real-time PCR were 21,611.5±8,294.7 and 75,997.5±39,709.3 (copies per 10 ng of total RNA), respectively.
The results indicate that P2X and P2Y receptor subtype mRNAs, except for P2X3, P2X6 and P2Y11 receptors, express in the rat gastric body muscle layer. Absolute quantitative real-time PCR analysis shows that P2Y1 and P2Y2 receptor mRNA expressions are significant higher than other P2Y receptor subtype mRNAs in the rat gastric body muscle layer, and the relative rank order of P2Y receptor subtype mRNA expression is P2Y2 > P2Y1 >> P2Y12 > P2Y6 = P2Y14 > P2Y13 > P2Y4 .
PartⅣP2Y1 and P2Y2 receptors detected using immunohisto- chemistry in smooth muscle cells of the rat gastric body
P2Y1 and P2Y2 receptor expressions in smooth muscle cells (SMCs) of the rat gastric body were detected using immunohistochemistry techniques, and we compared the distribution profile of P2Y1 and P2Y2 receptors between the gastric body and gastric fundus of the rat.
There was no specific immunoreactivity in the different layers of the gastric body and gastric fundus in control experimental preparations. In the rat gastric body and gastric fundus, P2Y1 receptor-immunoreactive (-ir) cells and P2Y2 receptor-ir cells were observed in the lamina propria, muscularis mucosae, longitudinal muscle (LM) layer, circular muscle (CM) layer and myenteric plexus. Strong immunostaining of P2Y1 and P2Y2 receptors was observed in the cytoplasm of gland cells and neurons, and in the nucleus of SMCs.
Percentages of P2Y1 receptor-ir and P2Y2 receptor-ir SMCs in the CM layer of the gastric body were 33.08%±9.93% and 45.17%±6.11% respectively, which were significantly higher than that in the LM layer and muscularis mucosae of the gastric body (P<0.01). The expression level of P2Y1 and P2Y2 receptor in SMCs of the CM layer, LM layer and muscularis mucosae of the gastric fundus was similar to the gastric body.
In CM and LM layers of the gastric body and fundus, percentages of P2Y2 receptor-ir SMCs were significantly higher than those of P2Y1 receptor-ir SMCs (P<0.05). The same phenomenon was detected in the muscularis mucosae SMCs of the gastric body.
Percentages of P2Y2 receptor-ir SMCs of the CM layer, LM layer and muscularis mucosae in the gastric body were significantly lower than that in the gastric fundus (P<0.05). Percentages of P2Y1 receptor-ir SMCs of the LM layer and muscularis mucosae in the gastric body were significantly lower than that in the gastric fundus (P<0.05). The expression of P2Y1 receptor in SMCs of the CM layer was the same in both gastric body and gastric fundus (P>0.05).
The results indicate that P2Y1 and P2Y2 receptors are highly expressed in nucleus of SMCs of the LM layer, CM layer and muscularis mucosae in the rat gastric body and gastric fundus. Both the receptors expressed in CM layer SMCs of the gastric body and gastric fundus are significantly higher than that in the SMCs of LM layer and muscularis mucosae. Both the receptors expressed in the SMCs of LM layer and muscularis mucosae in the gastric body are significantly lower than that in the gastric fundus. Moreover, in the rat gastric body and gastric fundus P2Y2 receptor expressions in SMCs of the LM layer and CM layer are significantly higher than P2Y1 receptor expressions.
Nucleotides producing muscle contraction in a concentration dependent manner are important mediators in circular muscle strips of the rat gastric body, and the pharmacological characteristics of circular muscle strips in the rat gastric body are different from that in gastric fundus.
There are several P2Y receptor subtypes involved in the contraction induced by nucleotides in circular smooth muscle of the rat gastric body. 2-MeSATP, ATP and ADP might act on a same P2Y receptor subtype regulating the contraction, and which is potentiated by a desensitization of P2Y2 and P2Y4 receptors.
P2Y1 and P2Y2 receptor mRNA expressions are significant higher than other P2Y receptor subtype mRNAs in smooth muscles of the rat gastric body, and the relative rank order of P2Y receptor subtype mRNA expression is P2Y2 > P2Y1 >> P2Y12 > P2Y6 = P2Y14 > P2Y13 > P2Y4 .
P2Y1 and P2Y2 receptors expressed in SMCs of the circular muscle layer are significantly higher than that of the longitudinal muscle layer in the rat gastric body and gastric fundus. Both the receptors expressed in SMCs of the longitudinal muscle layer in the rat gastric body are significantly lower than that in the rat gastric fundus. Moreover, in the rat gastric body and gastric fundus, P2Y2 receptor expression in SMCs of the circular and longitudinal muscle layers is higher than P2Y1 receptor expression.
引文
1 Otsuguro K, Ito S, Ohta T, et al. Influence of purines and pyrimidines on circular muscle of the rat proximal stomach. Eur J Pharmacol, 1996, 317(1): 97~105
2王秒,赵丁,赵庆华等.腺苷三磷酸对大鼠离体胃平滑肌运动的影响.中国药理学通报, 2005: 21(3): 351~355
3周吕.胃运动的功能//周吕,柯美云.胃肠动力学:基础与临床.第1版.北京:科学出版社, 1999: 509~527
4柯美云,谷成明,姜玉新等.?消化不良患者的胃幽门十二指肠运动协调性研究.中国医学科学院学报, 2000, 22(3): 241~245
5 Parkman HP, Harris AD, Krevsky B, et al. Gastroduodenal motility and dismotility: an update on techniques available for evaluation. Am J Gastroenterol, 1995, 90(6): 869~892
6 Tack J, Coulie B, Wilmer A, et al. Influence of sumatriptan on gastric fundus tone and on the perception of gastric distension in man. Gut, 2000, 46(4): 468~473
7 Marzio L, Cappello G, Grossi L, et al. Effect of the 5-HT3 receptor antagonist, ondansetron, on gastric size in dyspeptic patients with impairedgastric accommodation. Dig Liver Dis, 2008, 40(3): 188~193
8 Haas HL, Sergeeva OA, Selbach O. Histamine in the nervous system. Physiol Rev, 2008, 88(3): 1183~1241
9 Milenov K, Golenhofen K. Differentiated contractile responses of gastric smooth muscle to substance P. Pflügers Archiv, 1983, 397(1): 29~34
10 Matharu MS, Hollingsworth M. Purinoceptors mediating relaxation and spasm in the rat gastric fundus. Br J Pharmacol, 1992, 106(2): 395~403
11 Ren LM, Zhang M. Distribution of functional P2X1-like receptor in rabbit isolated regional arteries. Acta Pharmacol Sin, 2002, 23(8): 721~726
12 Hotta S, Morimura K, Ohya S, et al.Ryanodine receptor type 2 deficiency changes excitation–contraction coupling and membrane potential in urinary bladder smooth muscle. J Physiol, 2007, 582(2): 489~506
13 Gallego D, Hernández P, ClavéP, et al. P2Y1 receptors mediate inhibitory purinergic neuromuscular transmission in the human colon. Am J Physiol, 2006, 291(4): G584~G594
14 Zhao D, Ren LM. Non-adrenergic inhibition at prejunctional sites by agmatine of purinergic vasoconstriction in rabbit saphenous artery. Neuropharmacol, 2005, 48(4): 597~606
15 Guo ZD, Li Z, Ruan Y, et al. Distribution of muscarinic receptors of different of affinities in smooth muscle of human stomach. Acta Pharmacol Sin, 1991; 12(2): 180~183
16 Kitazawa T, Hashiba K, Cao J, et al. Functional roles of muscarinic M2 and M3 receptors in mouse stomach motility: Studies with muscarinic receptor knockout mice. Eur J Pharmacol, 2007; 554(2-3): 212~222
17 Venkova K, Milne A, Krier J. Contractions mediated byα1-adrenoceptors and P2-purinoceptors in a cat colon circular muscle. Br J Pharmacol, 1994, 112(4): 1237~1243
18 Giaroni C, Knight GE, Ruan H-Z, et al. P2 receptors in the murine gastrointestinal tract. Neuropharmacol, 2002, 43(8): 1313~1323
19 Burnstock G. Pathophysiology and therapeutic potential of purinergic signaling. Pharmacol Rev, 2006; 58(1): 58~86
20 Fredholm BB, Abbracchio MP, Burnstock G, et al. Nomenclature and classification of purinoceptors. Pharmacol Rev, 1994; 46(2): 143~156
21 King BF, Knowles ID, Burnstock G, et al. Investigation of the effects of P2 purinoceptor ligands on the micturition reflex in female urethane-anaesthetized rats. Br J Pharmacol, 2004, 142(3): 519~530
22 Ishiguchi T, Takahashi T, Itoh H, et al. Nitrergic and purinergic regulation of the pylorus. Am J Physiol, 2000, 279: G740~ G747
1 Mule F, Naccari D, Serio R. Evidence for the presence of P2y and P2x receptors with different functions in mouse stomach. Eur J Pharmacol, 2005, 513(1-2): 135~140
2 Otsuguro K, Ito S, Ohta T, et al. Influence of purines and pyrimidines on circular muscle of the rat proximal stomach. Eur J Pharmacol, 1996, 317(1): 97~105
3王秒,任雷鸣. P2Y受体介导大鼠胃体环行肌收缩反应的药理学特点.药学学报, 2009, 44(5): 473~479
4 Otsuguro K, Ohta T, Ito S, et al. Two types of relaxation-mediating P2 receptors in rat gastric circular muscle. Jpn J Pharmacol, 1998, 78(2): 209~215
5 Dunn PM,Blakeley AG. Suramin: a reversible P2-purinoceptor antagonist in the mouse vas deferens. Br J Pharmacol, 1988, 93(2): 243~245
6 Kennedy C. P1- and P2-purinoceptor subtypes-an update. Arch Int Pharmacodyn Ther, 1990, 303(1): 30~50
7 Kugelgen von I. Pharmacological profiles of cloned mammalian P2Y-receptor subtypes. Pharmacol Ther, 2006, 110(3): 415~432
8 Kugelgen von I, Wetter A. Molecular pharmacology of P2Y-receptors. Naunyn Schmiedebergs Arch Pharmacol, 2000, 362(4-5): 310~323
9 McDonnell B, Hamilton R, Fong M, et al. Functional evidence for purinergic inhibitory neuromuscular transmission in the mouse internal anal sphincter. Am J Physiol Gastrointest Liver Physiol, 2008, 294:G1041~1051
10 Kindig AE, Hayes SG, Hanna RL, et al. P2 antagonist PPADS attenuates responses of thin fiber afferents to static contraction and tendon stretch. Am J Physiol Heart Circ Physiol, 2006, 290: H1214~1219
11 Shehnaz D, Torres B, Balboa M, et al. Pyridoxal-phosphate-6- azophenyl-29,49-disulfonate(PPADS), a Putative P2Y1 Receptor Antagonist, Blocks Signaling at a Site Distal to the Receptor in Madin-Darby Canine Kidney-D1 Cells. J Pharmacol Exp Ther, 2000, 292(1): 346~350
12 Brown C, Tanna B, Boarder MR. PPADS: An antagonist at endothelial P2Y-purinoceptors but not P2U-purinoceptors. Br J Pharmacol, 1995, 116(5): 2413~2416
13 Ishiguchi T, Takahashi T, Itoh H, et al. Nitrergic and purinergic regulation of the pylorus. Am J Physiol, 2000, 279: G740~G747
14 Giaroni C, Knight GE, Ruan HZ, et al. P2 receptors in the murine gastrointestinal tract. Neuropharmacol, 2002, 43(8): 1313~1323
15 Giaroni C , Knight GE, Zanetti E, et al. Postnatal development of P2 receptors in the murine gastrointestinal tract. Neuropharmacol, 2006, 50(6): 690~704
16 Burnstock G. Pathophysiology and therapeutic potential of purinergic signaling. Pharmacol Rev, 2006; 58(1): 58~86
17 Fredholm BB, Abbracchio MP, Burnstock G, et al. Nomenclature and classification of purinoceptors. Pharmacol Rev, 1994, 46(2): 143~156
18 Bogdanov YD, Wildman SS, Clements MP, et al. Molecular cloning and characterization of rat P2Y4 nucleotide receptor. Br J Pharmacol, 1998, 124(3): 428~430
19 Homolya L, Steinberg TH, Boucher RC. Cell to cell communication in response to mechanical stress via bilateral release of ATP and UTP in polarized epithelia. J Cell Biol, 2000, 150(6): 1349~1359.
20 Clarke LL, Harline MC, Otero MA, et al. Desensitization of P2Y2 receptor-activated transepithelial anion secretion. Am J Physiol CellPhysiol, 1999, 276: C777~787
21 Windscheif U, Pfaff O, Ziganshin AU, et al. Inhibitory action of PPADS on relaxant responses to adenine nucleotides or electrical field stimulation in guinea-pig taenia coli and rat duodenum. Br J Pharmacol, 1995, 115(8): 1509~1517
22 Ralevic V, Burnstock G. Discrimination by PPADS between endothelial P2Y- and P2U-purinoceptors in the rat isolated mesenteric arterial bed. Br J Pharmacol, 1996, 118(2): 428~434
23 Rayment SJ, Latif ML, Ralevic V. Evidence for the expression of multiple uracil nucleotide-stimulated P2 receptors coupled to smooth muscle contraction in porcine isolated arteries. Br J Pharmacol, 2007, 150(5): 604~612
1 Crowe R, Burnstock G. Comparative studies of quinacrine-positive neurons in the myenteric plexus of stomach and intestine of guinea-pig, rabbit and rat. Cell Tissue Res, 1981, 221(1): 93~107
2 Belai A, Burnstock G. Evidence for the coexistence of ATP and nitric oxide in non-adrenergic, non-cholinergic (NANC) inhibitory neurones in the rat ileum, colon and annococcygeus muscle. Cell Tissue Res, 1994, 278(1): 197~200
3 Burnstock G, Cocks T, Kasakov L, et al. Direct evidence for ATP release from non-adrenergic, non-cholinergic (“purinergic”) nerves in the guinea-pig taenia coli and bladder. Eur J Pharmacol, 1978, 49(2): 145~149
4 McConalogue K, Todorov L, Furness JB, et al. Direct measurement of the release of ATP and its major metabolites from the nerve fibres of the guinea-pig taenia coli. Clin Exp Pharmacol Physiol, 1996, 23(9): 807~812
5 Burnstock G. Purinergic signalling in gut. Abbracchio MP, Williams M(Eds), Handbook of Experimental Pharmacology: Purinergic and Pyrimidinergic Signalling, Berlin: Springer, 2001, 151(2): 141~238
6 Misra S, Murthy KS, Zhou H, et al. Coexpression of Y1, Y2, and Y4 receptors in smooth muscle coupled to distinct signaling pathway. J Pharmacol Exp Ther, 2004, 311(12): 1154~1162
7 Suarez-Huerta N, Pouillon V, Boeynaems J. Molecular cloning and characterization of the mouse P2Y4 nucleotide receptor. Eur J Pharmacol, 2001, 416(3): 197~202
8 Bassil AK, Bourdu S, Townson KA, et al. UDP-glucose modulates gastric function through P2Y14 receptor-dependent and -independent mechanisms. Am J Physiol Gastrointest Liver Physiol, 2009, 296(4): G923~G930
9 Xiang Z, Burnstock G. Development of nerves expressing P2X3 receptors in the myenteric plexus of rat stomach. Histochem Cell Biol 2004, 122(2):111~119
10 Castelucci P, Robbins HL, Furness JB. P2X2 purine receptor immunoreactivity of intraganglionic laminar endings in the mouse gastrointestinal tract. Cell Tissue Res, 2003, 312(2): 167~174
11 Hu RG, Suzuki-Kerr H, Webb KF, et al. Molecular and functional mapping of regional differences in P2Y receptor expression in the rat lens. Exp Eye Res, 2008, 87(2): 137~146
12 Ma B, Yu LH, Fan J, et al. Pharmacological properties of P2 receptors on rat otic parasympathetic ganglion neurons. Life Sci, 2008, 83(5-6): 185~191
13 Carneiro-Ramos MS, Diniz GP, Almeida J, et al. Cardiac angiotensin II type I and type II receptors are increased in rats submitted to experimental hypothyroidism. J Physiol, 2007, 583(1): 213~223
14 Wittwer CT, Herrmann MG, Moss AA, et al. Continuous fluorescence monitoring of rapid cycle DNA amplification. Biotechniques, 1997, 22(1): 130~131, 134~138
15 Bustin SA. Absolute quantification of mRNA using real-time reverse transcription polymerase chain reaction assays. J Mol Endocrinol, 2000, 25(2): 169~193
16张传海,刘秋英,金琳等. nm23-H1基因实时荧光定量PCR标准质粒的构建.中国卫生检验杂志, 2007, 17(4): 604~615
17 Morrison TB, Weis JJ, Writtwer CT. Quantification of low-copy transcripts by continuous SYBR Green 1 monitoring during amplification. Biotechniques, 1998, 24(4): 954~962.
18 O’reilly BA, Kosaka AH, Chang TK, et al. A quantitative analysis of purinoceptor expression in human fetal and adults bladder. J Urol, 2001, 165(5), 1730~1734
19黄留玉. PCR最新技术原理、方法及应用(第1版).北京:化学工业出版社, 2005: 139~157
20 Wong ML, Medrano JF. Real-time PCR for mRNA quantitation. Biotechniques, 2005, 39 (1) : 75~85
21 Neuvians TP, Gashaw I, Sauer CG, et al. Standardization strategy for quantitative PCR in human seminoma and normal testis. J Biotechnol, 2005, 117(2): 163~171
1 Koshiba M, Apasov S, Sverdlov V, et al. Transient up-regulation of P2Y2 nucleotide receptor mRNA expression is an immediate early gene response in activated thymocytes. PNAS, 1997, 94(3): 831~836
2 Maaser K, Hopfner M, Kap H, et al. Extracellular nucleotides inhibits growth of human oesophageal cancer cells via P2Y(2)-receptors. Br J Cancer, 2002, 86(4): 636~644
3 Giaroni C, Knight GE, Ruan HZ, et al. P2 receptors in the murine gastrointestinal tract. Neuropharmacol, 2002, 43(8): 1313~1323
4 Giaroni C, Knight GE, Zanetti E, et al. Postnatal development of P2 receptors in the murine gastrointestinal tract. Neuropharmacol, 2006, 50(6): 690~704
5 Van Crombruggen K, Van Nassauw L, Timmermans JP, et al. Inhibitory purinergic P2 receptor characterisation in rat distal colon. Neuropharmacol, 2007, 53(2): 257~271
6 Gallego D, Herna′ndez P, Clave′P, et al. P2Y1 receptors mediate inhibitory purinergic neuromuscular transmission in the human colon. Am J Physiol Gastrointest Liver Physiol, 2006, 291(4): G584~G594
7 King BF, Townsend-Nicholson A. Involvement of P2Y1 and P2Y11 Purinoceptors in Parasympathetic Inhibition of Colonic Smooth Muscle. J Pharmacol Exp Ther, 2008, 324(3): 1055~1063
8 Xiang Z, Burnstock G. Distribution of P2Y2 receptors in the guinea pig enteric nervous system and its coexistence with P2X2 and P2X3 receptors, neuropeptide Y, nitricoxide synthase and calretinin. Histochem Cell Biol, 2005, 124(5):379~390
9 Xiang Z, Burnstock G. Distribution of P2Y6 and P2Y12 receptor: their colocalization with calbindin, calretinin and nitric oxide synthase in the guinea pig enteric nervous system. Histochem Cell Biol, 2006, 125(4): 327~336
10 Van Nassauw L, Costagliola A, Van Op den Bosch J, et al. Region-specific distribution of the P2Y4 receptor in enteric glial cells and interstitial cells of Cajal within the guinea-pig gastrointestinal tract. Auton Neurosci, 2006, 126-127(6): 299~306
11 Chopra B, Gever J, Barrick SR, et al. Expression and function of rat urothelial P2Y receptors. Am J Physiol Renal Physiol, 2008, 294: F821~F829
12 Matos JE, Sorensen MV, Geyti CS, et al. Distal colonic Na(+) absorption inhibited by luminal P2Y(2) receptors. Pflugers Arch, 2007, 454(6): 977~987
13 Coutinho-Silva R, Parsons M, Robson T, et al. Changes in expression of P2 receptors in rat and mouse pancreas during development and ageing. Cell Tissue Res, 2001, 306(3): 373~383
14 Vallejo AI, Bo XN, Burnstock G. P2Y purinoceptors in gastric glandsmembranes. Eur J Pharmacol, 1996, 312(2): 209~214
15 Van Crombruggen K, Van Nassauw L, Timmermans JP, et al. Inhibitory purinergic P2 receptor characterisation in rat distal colon. Neuropharmacol, 2007, 53(2): 257~271
1 Fredholm BB, Abbracchio MP, Burnstock G, et al. Nomenclature and classification of purinoceptors. Pharmacol Rev, 1994, 46(2): 143~156
2 Gorlach A. Control of adenosine transport by hypoxia. Circ Res, 2005, 97(1): 1~3
3 Ralevie V, Burnstock G. Receptors for pufines and pyrimidines. Pharmacol Rev, 1998, 50(3): 413~492.
4 Scholl DJ, Wells JN. Serine and alamine mutagenesis of the nine native cysterine residues of the human A1 adenosine receptors. Biochem Pharmacol, 2000, 60(11): 1647~1654
5 Dixon AK, Gubitz AK, Sirinathsinghji DJ, et al. Tissue distribution of adenosine receptor mRNAs in the rat. Br J Pharmacol, 1996, 118(6): 1461~1468
6 Brambilla D, Chapman D, Greene R. Adenosine mediation of presynaptic feedback inhibition of glutamate release. Neuron, 2005, 46(2): 275~283
7 Andoh T, Ishiwa D, Kamiya Y, et al. A1 adenosine receptor-mediated modulation of neuronal ATP-sensitive K channels in rat substantia nigra. Brain Res, 2006, 1124(1): 55~61
8 Barros RC, Branco LG, Cárnio EC. Respiratory and body temperature modulation by adenosine A1 receptors in the anteroventral preoptic region during normoxia and hypoxia. Respir Physiol Neurobiol, 2006, 153(2): 115~125
9 Ashton KJ, Peart JN,Morrison RR, et al. Genetic modulation of adenosine recep tor function and adenosine handling in murine hearts: insights and issues. J Mol Cell Cardiol, 2007, 42(4): 693~705
10 Sitkovsky M, Lukashev D. Regulation of immune cells by local tissue oxygen tension: HIF1αand adenosine receptors. Nat Rev Immunol, 2005, 5(9): 712~721
11 Sun D, Samuelson LC, Yang T, et al. Mediation of tubuloglomerular feedback by adenosine: Evidence from mice lacking adenosine 1 receptors. PNAS, 2001, 98(17): 9983~9988
12 Dhalla AK, Wong MY, Voshol PJ, et al. A1 adenosine receptor partial agonist lowers plasma FFA and improves insulin resistance induced by high fat diet in rodents. Am J Physiol Endocrinol Metab, 2007, 292(5): E1358~E1363
13 Ferre S, Fredholm BB, Morelli M, et al. Adenosine-dopamine receptor-receptor interaction as an intergrative mechanism in the basal ganglia. Trends Neurosci, 1997, 20(10): 482~487
14 Ongini E, Fredholm BB. Pharmacology of adenosine A2A receptors. Trends Pharmacol Sci, 1996, 17(10): 364~372
15 Urade Y, Eguchi N , Qu WM, et al. Minireview: Sleep regulation in adenosine A2A receptor-deficient mice. Neurology, 2003, 61 (Suppl 6): S94~S96
16 Moore KA, Nicoll RA, Schmitz D. Adenosine gates synaptic plasticity at hippocampal mossy fiber synapses. PNAS, 2003, 100(24): 14397~14402
17 Hamilton SP, Slager SL, De Leon AB, et al. Evidence for genetic linkage between a polymorphism in the adenosine 2A receptor and panic disorder. Neuropsychopharmacol, 2004, 9(3): 558~565
18 Kaster MP, Rosa AO, Rosso MM, et al. Adenosine administrationproduces an antidepressant-like effect in mice: evidence for the involvement of A1 and A2A receptors. Neurosci Lett, 2004, 355(1): 21~24
19 Yu L, Huang Z, Mariani J, et al. Selective inactivation or reconstitution of adenosine A2A receptors in bone marrow cells reveals their significant contribution to the development of ischemic brain injury. Nat Med, 2004, 10(10): 1081~1087
20 HaskóG, Pacher P. A2A receptors in inflammation and injury: lessons learned from transgenic animals. J Leukoc Biol, 2008, 83(3): 447~ 455.
21 Gazoni LM, Laubach VE, Mulloy DP, et al. Additive protection against lung ischemia-reperfusion injury by adenosine A2A receptor activation before procurement and during reperfusion. J Thorac Cardiovasc Surg, 2008, 135(1): 156~165
22李玮,朱佩芳,周元国.腺苷A2A受体的矛盾作用:保护还是加重损伤.生理科学进展, 2006, 37(3): 225~228
23 Chen JF, Pedata F. Modulation of ischemic brain injury and neuro-inflammation by adenosine A2A receptors. Curr Pharm Des, 2008, 14(15): 1490~1499
24 Thiel M, Caldwell CC, Sitkovsky MV. The critical role of adenosine A2A recep tors in downregulation of inflammation and immunity in the pathogenesis of infectious diseases. Microbes Infect, 2003, 5(6): 515~526
25 Duffy SM, Cruse G, Brightling CE, et al. Adenosine closes the K+ channel KCa3.1 in human lungmast cells and inhibits theirmigration via the adenosine A2A receptor. Eur J Immunol, 2007, 37(6): 1653~1662
26 Beukers MW, Dulk HD, Tilbury Erica WV, et al. Why are A2B receptor low-affinity adenosine recptors? Mutation of Asn273 to Tyr increases affinity of human A2B recptors for 2-(1-Hexynl) adeonosine. Mol Pharmacol, 2000, 58(6): 1349~1356
27 Vickram R, Gary LS, Michael AB, et al. The A3 adenosine receptor is the unique adenosine receptor which facilitates release of allergic mediators in mast cells. J Biol Chem, 1993, 268(23): 16887~16890
28 Linden J. Cloned adenosine A3 receptors: pharmacological properties,species differences and receptor functions. Trends Pharmacol Sci, 1994, 15(8): 298~306
29 Carruthers AM, Fozard JR. Effect of pertussis toxin treatment on the putative adenosine A3 receptor-mediated hypotensive response in the rat. Eur J Pharmacol, 1993, 250(1): 185~188
30 Maggirwar. Adenosine acts as an endogenous activator of the cellular antioxidant defense system. Biochem Biophys Res Commun, 1994, 201(2): 508~515
31 Gever JR, Cockayne DA, Dillon MP, et al. Pharmacology of P2X channels. Pflugers Arch, 2006, 452(5): 513~537
32 Vial C, Roberts JA, Evans RJ. Molecular properties of ATP-gated P2X receptorion channels. Trends Pharmacol Sci, 2004, 25(9): 487~493
33 North RA , Surprenant A. Pharmacology of cloned P2X receptors. Annu Rev Pharmacol Toxicol, 2000, 40: 563~580
34 Giniatullin R, Sokolova E, Nistri A. Modulation of P2X3 receptors by Mg2+ on rat DRG neurons in culture. Neuropharmacol, 2003, 44 (1): 132~140
35 Schulz A, Schoneberg T. The structural evolution of a P2Y2 like G-protein-coupled receptor. J Biol Chem, 2003, 278(37): 35531~35541
36 Abbracchio MP, Boeynaems JM, Barnard EA, et al. Characterization of the UDP-glucose receptor (renamed here the P2Y14 receptor) adds diversity to the P2Y receptor family. Trends Pharmacol Sci, 2003, 24(2): 52~55.
37 Ding Z, Tuluc F, Bandivadekar KR, et al. Arg333 and Arg334 in the COOH terminus of the human P2Y1 receptor are crucial for Gq coupling. Am J Physiol Cell Physiol, 2005, 288: C559~C567
38 Burnstock G, Campbell G, Bennett M, et al. Inhibition of the smooth muscle of the taenia coli. Nature, 1963, 200(9): 581~582
39 Burnstock G, Campbell G, Rand MJ. Innervation of the guinea-pig taenia coli: are there intrinsic inhibitory nerves which are distinct from sympathetic nerves? Int J Neuropharmacol, 1964, 3:163~166
40 Martinson J, Muren A. Excitatory and inhibitory effects of vagus stimulation on gastric motility in the cat. Acta Physiol, 1963, 57: 309~316
41 Burnstock G, Campbell G, Satchell D, et al. Evidence that adenosine triphosphate or a related nucleotide is the transmitter substance released by nonadrenergic inhibitory nerves in the gut. Br J Pharmacol, 1970, 40(4): 668~688
42 Burnstock G. Purinergic nerves. Pharmacol Rev, 1972, 24: 509~581
43 Burnstock G. Purinoceptors: ontogeny and phylogeny. Drug Dev Res, 1996, 39: 204~242
44 Burnstock G. Do some nerve cells release more than one transmitter? Neurosci, 1976, 1(4): 239~248
45 Kasakov L, Burnstock G. The use of the slowly degradable analog,α,β-methyleneATP, to produce desensitization of the P2-purinoceptor: effect on non-adrenergic, non-cholinergic responses of the guinea-pig urinary bladder. Eur J Pharmacol, 1982, 86(2): 291~294
46 Fedan JS, Hogaboom GK, O'Donnell, et al. Contributions by purines to the neurogenic response of the vas deferens of the guinea-pig. Eur J Pharmacol, 1981, 69(1): 41~53
47 Langer SZ, Pinto JEB. Possible involvement of a transmitter different from norepinephrine in residual responses to nerve stimulation of cat nicitating membrane after pretreatment with reserpine. J Pharmacol Exp Ther, 1976, 196(3): 697~713
48 Burnstock G, Warland JJ. A pharmacological study of the rabbit saphenous artery in vitro: a vessel with a large purinergic contractile response to sympathetic nerve stimulation. Br J Pharmacol, 1987, 90(1): 111~120
49 Burnstock G. Noradrenaline and ATP: cotransmitters and neuromodulators. J Physiol Pharmacol, 1995, 46(4): 365~384
50 Burnstock G. Purinergic signalling in lower urinary tract// Abbracchio MP, Williams M(Eds), Handbook of Experimental Pharmacology: Purinergic and Pyrimidinergic Signalling, Berlin: Springer, 2001,151: 423~515
51 Burnstock G. Purinergic signalling in gut//Abbracchio MP, Williams M(Eds), Handbook of Experimental Pharmacology: Purinergic and Pyrimidinergic Signalling, Berlin: Springer, 2001,151(2): 141~238
52 Burnstock G. Cotransmission. Curr Opin Pharmacol, 2004, 4(1): 47~52
53 Yamamoto K, Sokabe T, Ohura N, et al. Endogenously released ATP mediates shear stress-induced Ca2+ influx into pulmonary artery endothelial cells. Am J Physiol Heart Circ Physiol, 2003, 285: H793~H803
54 Aleu J, Martin-Satue M, Navarro P, et al. Release of ATP induced by hypertonic solutions in Xenopus oocytes. J Physiol, 2003, 547(1): 209~219
55 Schwiebert EM, Egan ME, Hwang TH, et al. CFTR regulates out-wardly rectifying chloride channels through an autocrine mechanism involving ATP. Cell, 1995, 81(7): 1063~1073
56 Braunstein GM, Roman RM, Clancy JP, et al. Cystic fibrosis trans-membrane conductance regulator facilitates ATP release by stimulating a separate ATP release channel for autocrine control of cell volume regulation. J Biol Chem, 2001, 276(9): 6621~6630
57 Zimmermann H. Ectonucleotidases: some recent developments and a note on nomenclature. Drug Dev Res, 2001, 52(1): 44~56
58 Burnstock G. Purinergic receptors in the nervous system// Schwiebert EM(Eds). Current Topics in Membranes: Purinergic Receptors and Signalling , 2003, 54: 307~368, Academic Press
59 Wang X, Arcuino G, Takahiro T, et al. P2X7 receptor inhibition improves recovery after spinal cord injury. Nat Med, 2004, 10(8): 821~827
60 Davalos D, Grutzendler J , Yang G, et al. ATP mediates rapid microglial response to local brain injury in vivo. Nat Neurosci, 2005, 8(6): 752~758
61 Haynes SE, Hollopeter G, Yang G, et al. The P2Y12 receptor regulates microglial activation by extracellular nucleotides. Nat Neurosci, 2006, 9(12): 1512~1519
62 Koizumi S, Shigemoto-mogami Y, Nasu-Tada K, et al. UDP acting P2Y6receptors is a mediator of microglial phagocytosis. Nature, 2007, 446(7139): 1091~1095
63 Khakh BS, Bumstock G, Kermedy C, et a1. Intemational Union of Phallnacology. XXIV. Current status of the nomenclature and properties of P2X receptors and their subunits. Pharmacol Rev. 2001, 53(1): 107~l18
64 Burnstock G. A unifying purinergic hypothesis for the initiation of pain. Lancet, 1996, 347(9015): 1604~1605
65 Tsuda M, Toyomitsu E, Komatsu T, et al. Lyn tyrosine kinase is required for P2X4 receptor up regulation and neuropathic pain after peripheral nerve injury. Glia, 2008, 56(5): 579~585
66 Yerxa BR. Therapeutic use of nucleotides in respiratory and ophthalmic diseases. Drug Dev Res, 2001, 52, 196–201
67 Bailey MA, Turner C, Huscitharel A, et a1. P2Y receptors present in the native and isolated rat glomerulus. Nephron Physiol, 2004, 96(3): 79~90
68 Burnstock GE, Knight GE. Cellular distribution and functions of P2 receptor subtypes in different systems. Int Rev Cytol, 2004, 240(1): 31~304
69 Burnstock G. Purinergic signalling and vascular cell proliferation and death. Arterio Thromb Vasc Biol, 2002, 22(3): 364~373
70 Wheeler-Schilling TH, Marquordt K, Kohler K, et al. Identification of purinergic receptors in retinal ganglion cells. Brain Res Mol Brain Res, 2001, 92(1-2): 177~180
71 Pintor J, Peral A, Pelaez T, et al. Nucleotides and dinucleotides in ocular physiology: New possibilities of nucleotides as therapeutic agents in the eye. Drug Dev Res, 2003, 59: 136~145
72 Sugiyama T, Kobayashi M, Kawamura H, et al. Enhancement of P2X7-induced pore formation and apoptosis: an early effect of diabetes on the retinal microvasculature. Invest Ophthalmol Vis Sci, 2004, 45(3): 1026~1032
73 Housley GD. Physiological effects of extracellular nucleotides in the inner ear. Clin Exp Pharmacol Physiol, 2000, 27(8): 575~580
74 Wang JC, Raybould NP, Luo L, et al. Noise induces up-regulation of P2X2 receptor subunit of ATP-gated ion channels in the rat cochlea. Neuroreport, 2003, 14(6): 817~823
75 Gale JE, Piazza V, Ciubotaru CD, et al. A mechanism for sensing noise damage in the inner ear. Curr Biol, 2004, 14(6): 526~529
76 Gayle S, Burnstock G. Immunolocalization of P2X and P2Y nucleotide receptors in the rat nasal mucosa. Cell Tissue Res, 2005, 319(1): 27~36
77 Finger TE, Danilova V, Barrows J, et al. ATP signaling is crucial for communication from taste buds to gustatory nerves. Science, 2005, 310(12): 1495~1499
78 Galligan JJ, North RA. Pharmacology and function of nicotinic acetylcholine and P2X receptors in the enteric nervous system. Neurogastroenterol Motil,2004,16(Suppl 1): 64~70
79 Castelucci P, Robbins HI, Poole DP, et al. The distribution of purine P2X2 receptors in the guinea-pig enteric nervous system. Histochem Cell Biol, 2002, 117(5): 415~422
80 Ruan HZ,Burnstock G. The distribution of P2X5 purinergic receptors in the enteric nervous system of mouse. Cell Tissue Res, 2005, 319(2): 191~200
81 Christofi FL, Guan Z, Wood JD, et al. Purinergic Ca2+ signaling in myenteric neurons via P2 purinoceptors. Am J Physiol, 1997, 272: G463~G473
82 Gourine AV, Laudet E, Dale N, et al. ATP is a mediator of chemosensory transduction in the central nervous system. Nature, 2005, 436(7047): 108~111