多沙唑嗪光学异构体降低动物膀胱排尿压的作用机制研究
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摘要
良性前列腺增生(benign prostatic hyperplasia,BPH)是老年男性常见病,常导致下尿路症状(lower urinary tract symptoms,LUTS),严重影响患者的生活质量。长效α1受体阻断药消旋多沙唑嗪(racemic-doxazosin, rac-DOX)可阻断下尿路α1受体,降低患者排尿阻力,缓解BPH/LUTS症状,是临床治疗BPH/LUTS的常用药物。但是,rac-DOX常引起头晕、头痛和低血压等不良反应。因此,应用α1受体阻断药治疗BPH/LUTS时,应考虑药物的“尿路选择性”。rac-DOX由光学异构体S-多沙唑嗪(S-doxazosin, S-DOX)和R-多沙唑嗪(R-doxazosin, R-DOX)组成。药理学研究结果表明,rac-DOX、S-DOX和R-DOX阻断人前列腺组织中α1受体的pA2值相同。本室研究发现,S-DOX降低动物血压和阻断血管α?受体的作用明显弱于rac-DOX和R-DOX,但S-DOX降低膀胱排尿压的作用与rac-DOX相同。为了分析药物降低动物膀胱排尿压的作用机制,我们采用兔离体膀胱、前列腺及尿道组织,观察了多沙唑嗪及其光学异构体的作用,同时利用老年小鼠观察多沙唑嗪及其光学异构体对前列腺细胞增殖与凋亡的影响。
第一部分多沙唑嗪光学异构体对兔离体膀胱逼尿肌的作用和作用机制
本研究观察了S-DOX、R-DOX和rac-DOX对兔离体膀胱逼尿肌收缩反应的影响。利用兔离体背侧和腹侧膀胱逼尿肌标本,分析多沙唑嗪及其光学异构体对药物或神经刺激诱发兔离体膀胱逼尿肌收缩反应的影响并分析其作用机制。
1背侧、腹侧膀胱逼尿肌对卡巴胆碱和苯肾上腺素诱发收缩反应的比较卡巴胆碱(0.01~30μmol?L-1)使背侧和腹侧膀胱逼尿肌产生浓度依赖性收缩反应,卡巴胆碱的浓度-反应曲线在两种标本上无显著性差异(P>0.05);与卡巴胆碱不同,苯肾上腺素(0.1~300μmol?L-1)仅使背侧膀胱逼尿肌产生浓度依赖性收缩反应,对腹侧膀胱逼尿肌无收缩作用。
2 S-DOX、R-DOX及rac-DOX对苯肾上腺素诱发兔背侧膀胱逼尿肌收缩反应的影响
苯肾上腺素使背侧膀胱逼尿肌产生浓度依赖性收缩反应,溶剂对苯肾上腺素诱发的兔背侧膀胱逼尿肌收缩反应无影响(P>0.05)。S-DOX、R-DOX和rac-DOX拮抗苯肾上腺素诱发兔背侧膀胱逼尿肌收缩反应的pKB值分别为7.44±0.19、7.39±0.14和7.38±0.30,三者之间无显著性差异(P>0.05)。
3 S-DOX、R-DOX及rac-DOX对电场刺激诱发兔背侧、腹侧膀胱逼尿肌收缩反应的影响
电场刺激诱发兔背侧和腹侧膀胱逼尿肌产生稳定的收缩反应,该收缩反应被0.1μmol?L-1的河豚毒素(TTX)完全阻断。S-DOX、R-DOX和rac-DOX均显著抑制电场刺激诱发的背侧膀胱逼尿肌收缩反应(P<0.01),且三者的抑制作用强度无显著性差异(P>0.05);但是,它们对电场刺激诱发的兔腹侧膀胱逼尿肌收缩反应无影响(P>0.05)。
上述结果表明,S-DOX、R-DOX和rac-DOX通过阻断膀胱逼尿肌α1受体显著抑制电场刺激诱发的兔背侧膀胱逼尿肌收缩反应,三者的抑制作用强度无显著性差异。
第二部分多沙唑嗪光学异构体对兔离体前列腺平滑肌收缩反应的影响
本研究观察了S-DOX、R-DOX和rac-DOX对兔离体前列腺收缩反应的影响。利用兔离体前列腺标本,分析多沙唑嗪及其光学异构体对药物或神经刺激诱发兔离体前列腺收缩反应的影响并分析其作用机制。
1 S-DOX、R-DOX及rac-DOX对苯肾上腺素诱发兔离体前列腺收缩反应的影响
苯肾上腺素使前列腺产生浓度依赖性收缩反应,溶剂对苯肾上腺素诱发的兔离体前列腺收缩反应无影响(P>0.05)。0.01μmol·L~(-1)和0.03μmol·L~(-1)的S-DOX、R-DOX或rac-DOX均可使苯肾上腺素量效曲线平行右移,Emax值不变。Schild plot分析结果表明,S-DOX、R-DOX或rac-DOX竞争性拮抗苯肾上腺素诱发的兔离体前列腺收缩反应,其pA2值分别为8.22±0.03、8.25±0.03和8.20±0.04,三者间无显著性差异(P>0.05)。S-DOX、R-DOX和rac-DOX拮抗苯肾上腺素诱发兔前列腺收缩反应的作用强度相同。
2 S-DOX、R-DOX及rac-DOX对电场刺激诱发兔离体前列腺收缩反应的影响
电场刺激诱发兔离体前列腺产生稳定的收缩反应,该收缩反应被0.1μmol·L~(-1)的TTX完全阻断。0.01μmol·L~(-1)或0.1μmol·L~(-1)的S-DOX、R-DOX和rac-DOX显著抑制电场刺激诱发的兔前列腺收缩反应(P<0.01),三者对电场刺激诱发兔离体前列腺收缩反应的抑制强度无显著性差异(P>0.05)。
3 Atr、Phent以及PPADS对电场刺激诱发兔离体前列腺收缩反应的影响
1μmol·L~(-1)Atr单独应用显著抑制电场刺激诱发的前列腺收缩反应,抑制百分率为62.6±11.7%(P<0.01),联合应用10μmol·L~(-1)的Phent后,二者完全抑制电场刺激诱发的前列腺收缩反应。单独给予10μmol·L~(-1)Phent可显著抑制电场刺激诱发的前列腺收缩反应,抑制百分率为96.7±9.3%(P<0.01),继续给予1μmol·L~(-1)Atr后,二者完全抑制电场刺激诱发的前列腺收缩反应。10μmol·L~(-1)PPADS对电场刺激诱发的前列腺收缩反应无明显影响(P>0.05)。TTX(0.1μmol·L~(-1))完全阻断该条件下电场刺激诱发的收缩反应。
上述结果表明,多沙唑嗪及其光学异构体显著抑制电场刺激诱发的兔离体前列腺收缩反应,其作用机制与药物阻断突出后α1受体有关,三者对兔离体前列腺α1受体的亲和力相同。
第三部分多沙唑嗪光学异构体对兔离体尿道环肌收缩功能的影响
制备兔离体膀胱颈和尿道环肌标本,记录药物或神经刺激诱发的收缩反应。观察多沙唑嗪及其光学异构体对苯肾上腺素诱发兔离体膀胱颈和尿道环肌收缩反应的影响;分析药物对电场刺激诱发兔离体尿道环肌收缩反应的作用及其作用机制。
1 S-DOX、R-DOX及rac-DOX对苯肾上腺素诱发兔膀胱颈和尿道环肌收缩反应的影响
苯肾上腺素使膀胱颈和各部位尿道环肌产生浓度依赖性收缩反应,溶剂对苯肾上腺素诱发的兔膀胱颈和尿道环肌收缩反应无影响(P>0.05)。0.03μmol·L~(-1)的S-DOX、R-DOX或rac-DOX拮抗苯肾上腺素诱发的兔离体膀胱颈收缩反应的pKB值分别为7.88±0.23、7.92±0.11和7.87±0.10,三者之间无显著性差异(P>0.05)。同样,在上段尿道、中段尿道和下段尿道,S-DOX、R-DOX和rac-DOX拮抗苯肾上腺素诱发各段尿道环肌收缩反应的pKB值亦无显著性差异(P>0.05)。
2 rac-DOX对电场刺激诱发兔膀胱颈和尿道环肌收缩反应的影响
溶剂对各标本的频率-反应曲线无明显影响(P>0.05)。不同浓度的rac-DOX(0.01、0.1和1μmol·L~(-1))对电场刺激诱发的兔离体膀胱颈和各段尿道环肌收缩反应无影响(P>0.05)。TTX(0.1μmol·L~(-1))完全阻断该条件下电场刺激诱发的收缩反应。
3 Pra、Atr及Ind对电场刺激诱发兔膀胱颈和尿道环肌收缩反应的影响
溶剂对各标本的频率-反应曲线无明显影响(P>0.05),TTX(0.1μmol·L~(-1))完全阻断电场刺激诱发的收缩反应。单独给予1μmol·L~(-1)的Atr显著抑制电场刺激诱发的兔膀胱颈收缩反应(P<0.05)。1μmol·L~(-1)的Pra对电场刺激诱发的膀胱颈收缩反应无显著影响;联合应用1μmol·L~(-1)的Atr后,抑制作用与单独给予Atr无显著性差异;与Pra和Atr联合用药组比,合用1μmol·L~(-1)的Ind进一步抑制了电场刺激诱发的兔离体膀胱颈收缩反应(P<0.01)。
在上段、中段以及下段尿道,1μmol·L~(-1)的Pra、Atr和Ind单独及联合应用均对电场刺激诱发的收缩反应无显著影响(P>0.05)。
4 Atr、Phent、Tub、尼古丁、苏拉明及TTX对电场刺激诱发兔尿道环肌收缩反应的影响
电场刺激诱发兔尿道环肌产生稳定的收缩反应。Atr(0.1~3μmol·L~(-1))和Phent(1~30μmol·L~(-1))对电场刺激诱发的兔尿道环肌收缩反应无影响。0.01μmol·L~(-1)和0.03μmol·L~(-1)的Tub显著抑制电场刺激诱发的收缩反应,抑制百分率分别为11.4±6.2%(P<0.05)和52.1±14.8%(P<0.01)。10μmol·L~(-1)的尼古丁显著抑制电场刺激诱发的收缩反应,抑制百分率为53.7±14.9%(P<0.01)。10、30、和100μmol·L~(-1)的苏拉明显著抑制电场刺激诱发的收缩反应,抑制百分率分别为22.5±3.5%、33.8±5.3%和54.2±5.9%。此外,0.1μmol·L~(-1)的Tub、30μmol·L~(-1)的尼古丁以及0.1μmol·L~(-1)的TTX尚能完全阻断电场刺激诱发的收缩反应。
5 CCh、Mox、Clo、Ado、ATP、UTP、5-HT和His对兔尿道环肌的作用
Mox(10μmol·L~(-1))和Clo(10μmol·L~(-1))可诱发尿道环肌产生较小收缩反应,标本最大收缩反应分别为0.20±0.12g和0.11±0.05g。CCh(30μmol·L~(-1))、Ado(30μmol·L~(-1))、ATP(10μmol·L~(-1))、UTP(30μmol·L~(-1))、5-HT(3μmol·L~(-1))和His(30μmol·L~(-1))对尿道标本均无收缩作用。
上述结果表明,S-DOX、R-DOX和rac-DOX通过阻断α1受体拮抗苯肾上腺素诱发的兔膀胱颈和尿道环肌收缩反应,三者阻断α1受体的作用相同。电场刺激诱发的兔尿道环肌收缩反应具有骨骼肌运动神经兴奋的特性,这一收缩成分同时易被尼古丁或苏拉明阻断。
第四部分多沙唑嗪光学异构体对老年小鼠前列腺增生的影响
实验采用老年小鼠长期(12周)灌胃给药法,观察了S-DOX、R-DOX和rac-DOX对老年小鼠前列腺体积、湿重、细胞增殖周期与凋亡百分率的影响。
1 S-DOX、R-DOX和rac-DOX对前列腺体积和湿重的影响与青年对照组相比,老年组小鼠前列腺体积指数无显著性差异,而S-DOX 6.0mg·kg-1组、R-DOX 6.0mg·kg-1组和R-DOX 0.6mg·kg-1组以及rac-DOX 6.0mg·kg-1组小鼠前列腺体指数显著减小(P<0.05)。
2 S-DOX、R-DOX和rac-DOX对小鼠前叶前列腺细胞增殖周期及凋亡的影响
与青年对照组相比,老年组小鼠前叶前列腺G0/G1期细胞周期百分率显著降低(P<0.01), G2/M期细胞周期百分率和增殖指数均显著增高(P<0.01);老年组小鼠前列腺细胞细胞凋亡率无明显改变。与老年组相比,S-DOX、R-DOX和rac-DOX老年小鼠前列腺细胞的增值指数和细胞周期无显著影响。
上述结果表明,长期(12周)灌胃给予S-DOX、R-DOX和rac-DOX,可能轻度降低老年小鼠前列腺细胞体积指数。
结论
S-DOX对心血管系统的抑制作用明显弱于R-DOX和rac-DOX,但是其降低膀胱排尿压的作用与同剂量rac-DOX相同。本研究证明,S-DOX降低动物膀胱排尿压的作用与其抑制去甲肾上腺素能神经兴奋所致的膀胱逼尿肌和前列腺平滑肌收缩反应有关;其作用机制是阻断了上述平滑肌组织的α1受体。S-DOX的上述作用及作用强度与R-DOX相同,说明兔膀胱逼尿肌和前列腺平滑肌细胞膜上的α1受体对多沙唑嗪光学异构体的亲和力相同。
S-DOX、R-DOX及rac-DOX可阻断兔尿道环肌α1受体,三者的作用强度相同;但是α1受体阻断药rac-DOX及哌唑嗪不影响电场刺激诱发的兔尿道环肌收缩反应。电场刺激诱发的兔尿道环肌收缩反应具有骨骼肌运动神经兴奋的特性,这一收缩成分同时易被尼古丁或苏拉明阻断。
Benign prostatic hyperplasia (BPH) often leads to lower urinary tract symptoms (LUTS). Patients with BPH/LUTS are suffering serious troubles on the quality of life. racemic-Doxazosin (rac-DOX),a long acting selectiveα1-adrenoceptor antagonist, is the current medication for the clinical treatment of BPH/LUTS. Several clinical studies have demonstrated that rac-DOX produces a decrease in urethral resistance and improve in lower urinary tract symptoms by blockingα1-adrenoceptor. However, rac-DOX is limited in the clinical application for BPH/LUTS due to its common adverse effects of dizziness, headache, and orthostatic hypotension. Therefore, the urinary tract selectivity should be necessarily considered whenα1-adrenoceptor antagonists are used for the BPH/LUTS patients.
rac-DOX is a mixture of S-doxazosin (S-DOX) and R-doxazosin (R-DOX). In a pharmacological study using the human isolated prostate tissue, S-DOX, R-DOX and rac-DOX all competitively antagonized the phenylephrine-induced contractile responses without significant differences in the pA2 values among S-DOX, R-DOX and rac-DOX. Recently, a study in our laboratory showed that the effects of S-DOX on animal blood pressure and onα1-adrenoceptors of the isolated rabbit blood vessels were significantly weaker than rac-DOX and R-DOX. However,the inhibitory effect by S-DOX on vesical micturition pressure was same to rac-DOX. To investigate the mechanisms of decreasing vesical micturition pressure by doxazosin and its enantiomers in animals, we observed the effects of S-DOX, R-DOX and rac-DOX on the contractile responses to drugs and electric field stimulation in the isolated rabbit urinary bladder detrusor, prostate and urethral strips, and the effects of S-DOX, R-DOX and rac-DOX on the prostate cell proliferation and apoptosis in the aged mouse.
PartⅠEffects of doxazosin and its enantiomers on the contractile responses in the isolated rabbit urinary detrusor strips
Contractile responses to drugs or electric field stimulation in the dorsal and ventral detrusor strips of the isolated rabbit urinary bladder were recorded to study the effects of S-DOX, R-DOX and rac-DOX on the contractile responses.
1 Contractile responses to carbachol and phenylephrine in the dorsal and ventral detrusor strips of the isolated rabbit urinary bladder
Carbachol produced contractile responses concentration-dependently in the dorsal and ventral detrusor strips, and the concentration-response curves for carbachol in the two kinds of strips were not significantly different from each other. Phenylephrine induced contractile responses in a concentration-dependent manner in the dorsal detrusor strips but not in the ventral detrusor strips.
2 Effects of doxazosin and its enantiomers on the contractile responses to phenylephrine in the dorsal detrusor strips of the isolated rabbit urinary bladder
Phenylephrine produced contractile responses concentration-dependently in the dorsal detrusor strips, and solvent had no effect on the contractile responses. The pKB values of S-DOX, R-DOX and rac-DOX at 1μmol·L~(-1) against phenylephrine in the dorsal detrusor strips of the isolated rabbit urinary bladder were 7.44±0.19, 7.39±0.14 and 7.38±0.30, respectively, and the pKB values of doxazosin and its enantiomers were not significantly different from each other (P>0.05).
3 Effects of doxazosin and its enantiomers on the contractile responses to electric field stimulation in the dorsal and ventral detrusor strips of the isolated rabbit urinary bladder
Electric field stimulation produced a steady contractile responses those were completely inhibited by tetrodotoxin (TTX) at 0.1μmol·L~(-1). S-DOX, R-DOX and rac-DOX significantly inhibited the contractile responses to electric field stimulation in the dorsal detrusor strips of the isolated rabbit urinary bladder (P<0.05), and their inhibitory effects were not significantly different from each other (P>0.05). However, S-DOX, R-DOX and rac-DOX did not affect the responses to electric field stimulation in the ventral detrusor strips.
These results indicate that S-DOX, R-DOX and rac-DOX significantly inhibit the contractile responses to electric field stimulation in the dorsal detrusor strips of the isolated rabbit urinary bladder via postsynapticα?1-adrenoceptors, and S-DOX inhibits the responses to the same extent as R-DOX and rac-DOX.
PartⅡEffects of doxazosin and its enantiomers on contractile responses in the isolated rabbit prostate strips
Contractile responses to drugs or electric field stimulation were recorded to study the effects of S-DOX, R-DOX and rac-DOX on the contractile responses in the isolated rabbit prostate strips.
1 Effects of doxazosin and its enantiomers on the contractile responses to phenylephrine in the isolated rabbit prostate strips
Phenylephrine produced contractile responses concentration-dependently in the prostate strips, and solvent did not affect the contractile responses. S-DOX, R-DOX and rac-DOX at 0.01μmol·L~(-1) and 0.03μmol·L~(-1) produced parallel shift of the cumulative concentration-response curves (CCRCs) for phenylephrine to the right in the isolated rabbit prostate, and the values of Emax of CCRCs for phenylephrine were not changed significantly by S-DOX, R-DOX and rac-DOX (P>0.05). The Schild plot analysis indicated that S-DOX, R-DOX and rac-DOX competitively antagonized the phenylephrine-induced contractile responses in the isolated rabbit prostate strips. The pA2 values of S-DOX, R-DOX and rac-DOX were 8.22±0.03, 8.25±0.03 and 8.20±0.04, respectively, and the pA2 values of three agents were not significantly different from each other (P>0.05).
2 Effects of doxazosin and its enantiomers on the contractile responses to electric field stimulation in the isolated rabbit prostate strips
Electric field stimulation produced a steady contractile responses, and the responses were completely inhibited by TTX at 0.1μmol·L~(-1). S-DOX, R-DOX and rac-DOX significantly inhibited the contractile responses to electric field stimulation in the isolated rabbit prostate strips (P<0.01), and S-DOX inhibited the responses to the same extent as R-DOX and rac-DOX.
3 Effects of atropine, phentolamine and PPADS on the contractile responses to electric field stimulation in the isolated rabbit prostate strips
Atropine (1μmol·L~(-1)) significantly reduced the contractile responses to electric field stimulation by 62.6±11.7%, and co-application with 10μmol·L~(-1) phetolamine abolished the contractile responses to electric field stimulation. Phetolamine (10μmol·L~(-1)) alone markedly reduced the contractile responses to electric field stimulation by 96.7±9.3%, and co-application with 1μmol·L~(-1) atropine abolished the contractile responses to electric field stimulation. PPADS (10μmol·L~(-1)) failed to affect the contraction induced by electric field stimulation in the isolated rabbit prostate strips (P>0.05). TTX (0.1μmol·L~(-1)) abolished the contractile responses.
These results indicate that doxazosin and its enantiomers significantly inhibit the contractile responses to electric field stimulation via postsynapticα1-adrenoceptors in the isolated rabbit prostate strips. The pA2 values of doxazosin and its enantiomers against phenylephrine are not significantly different from each other in the isolated rabbit prostate strips.
PartⅢEffects of doxazosin and its enantiomers on the contractile responses in the isolated rabbit urethral strips
In the urinary bladder neck strip and proximal, medial and distal urethral strips of the rabbit, the contractile responses to drugs and electric field stimulation were recorded. The effects of S-DOX, R-DOX and rac-DOX on the contractile responses to phenylephrine in the urinary bladdr neck strip and proximal, medial and distal urethral strips were observed. The pharmacological prolife of contractile responses to electric field stimulation in the urethral strips were investigated.
1 Effects of doxazosin and its enantiomers on the contractile responses to phenylephrine in the isolated rabbit urinary bladder neck strip and urethral strips
Phenylephrine produced contractile responses concentration-dependently in the isolated rabbit urinary bladder neck strip and the urethral strips, and solvent did not affect the contractile responses (P>0.05). The pKB values of S-DOX, R-DOX and rac-DOX against phenylephrine in the bladder neck were 7.88±0.23, 7.92±0.11 and 7.87±0.10, respectively, and the pKB values of the three agents were not significantly different from each other (P>0.05). The pKB values of doxazosin and its enantiomers against phenylephrine were not significantly different in the proximal, medial and distal urethral strips.
2 Effects of rac-DOX on the contractile responses to electric field stimulation in the isolated rabbit urinary bladder neck strip and proximal, medial and distal urethral strips
Solvent did not affect the frequency-response curves for electric field stimulation in the isolated rabbit urinary bladder neck strip and urethral strips. rac-DOX (0.01-1μmol·L~(-1)) was ineffective against the contractions induced by electric field stimulation in the isolated urinary bladder neck strip and proximal, medial and distal urethral strips, even though TTX at 0.1μmol·L~(-1) completely inhibited the responses.
3 Effects of prazosin, atropine and indomethacin on the contractile responses to electric field stimulation in the isolated rabbit urinary bladder neck strip and proximal, medial and distal urethral strips
Solvent did not affect the frequency-response curves for electric field stimulation in the isolated rabbit urinary bladder neck strip and urethral strips. Prazosin (1μmol·L~(-1)) was ineffective against the contractile responses to electric field stimulation (P>0.05), while atropine (1μmol·L~(-1)) significantly reduced the contractions (P<0.01). Co-application of prazosin and atropine caused an inhibition to the same extent as atropine alone (P>0.05). A treatment with the combination of prazosin, atropine and indomethacin decreased the contractile responses more potently than a treatment with a combination of prazosin and atropine (P<0.01). In the proximal, medial and distal urethral strips of the rabbit, prazosin, atropine or indomethacin did not affect the contractile responses to electric field stimulation, even though TTX at 0.1μmol·L~(-1) abolished the responses.
4 Effects of atropine, phentolamine, tubocurarine, nicotine, suramin and tetrodotoxin on the contractile responses to electric field stimulation in the isolated rabbit urethral strips
Electric field stimulation produced a steady contractile response in the isolated rabbit urethral strips. Atropine (0.1-3μmol·L~(-1)) or phentolamine (1-30μmol·L~(-1)) did not affect the contraction induced by electric field stimulation in the isolated rabbit urethral strips. Tubocurarine significantly inhibited the contraction at 0.01μmol·L~(-1) and 0.03μmol·L~(-1) by 11.4±6.2% and 52.1±14.8%, respectively. Nicotine at 10μmol·L~(-1) significantly inhibited the contraction by 53.7±14.9%. Suramin at 10, 30 and 100μmol·L~(-1) significantly inhibited the contraction by 22.5±3.5%, 33.8±5.3% and 54.2±5.9%, respectively.
Tubocurarine (0.1μmol·L~(-1)), Nicotine (30μmol·L~(-1)) and TTX (0.1μmol·L~(-1)) abolished the contractile responses to electric field stimulation in the isolated rabbit urethral strips. 5 Responses of the isolated rabbit urethral strips to carbachol, moxonidine, clonidine, adenosine, ATP, UTP, 5-HT and histamine
Moxonidine (10μmol·L~(-1)) and clonidine (10μmol·L~(-1)) produced small contractions in the isolated rabbit urethral strips, and the contractile force was 0.20±0.12g and 0.11±0.05g, respectively. Carbachol (30μmol·L-1), adenosine (30μmol·L~(-1)), ATP (10μmol·L~(-1)), UTP (30μmol·L~(-1)), 5-HT (3μmol·L~(-1)) and histamine (30μmol·L~(-1)) could not produce any contractile responses in the urethral strips.
These results indicate that doxazosin and its enantiomers antagonize the phenylephrine-induced contractile responses in the isolated rabbit urinary bladder neck strips and proximal, medial and distal urethral strips via postsynapticα1-adrenoceptors, and the pKB values of doxazosin and its enantiomers are not significantly different from each other.
Neurogenic contractile responses to electric field stimulation are motor neuron-evoked contractions with characteristics of highly sensitive to nicotine and suramine, and the neurogenic contractions are not affected by rac-DOX and prazosin.
PartⅣEffects of doxazosin and its enantiomers on the volume and cell proliferation of the prostate in aged mouse
The male mouse at 13 months of age was used to study the effects of long-term (12 weeks) intragastrical administration of doxazosin and its enantiomers on the volume and wet weight of the prostate, and the effects on the cell proliferation and apoptosis of the anterior lobe prostatic cells in aged mouse.
1 Effects of doxazosin and its enantiomers on the volume and wet weight of the prostate in aged mouse
In comparison with young mouse, the volume index of the prostate in aged mouse did not change significantly (P>0.05), but the volume index of the prostate in aged mouse treated with S-DOX (6.0mg·kg-1), R-DOX (6.0mg·kg-1), R-DOX (0.6mg·kg-1) or rac-DOX (6.0mg·kg-1) was significantly decreased (P<0.05).
2 Effects of doxazosin and its enantiomers on the cell cycle, proliferation and apoptosis of the anterior lobe cells of the prostate in aged mouse
In comparison with young mouse, the total G0/G1-phase of the cells of the anterior lobe of the prostate were significantly decreased (P<0.01), and the total G2/M-phase cells of the anterior lobe of the prostate and the proliferation index of the prostatic cells were significantly increased in aged mouse (P<0.01).
In comparison with aged mouse, the proliferation index and cell cycle distribution of the anterior prostatic lobe in aged mouse treated by S-DOX, R-DOX and rac-DOX did not change significantly (P>0.05).
These results indicate that doxazosin and its enantiomers slightly decrease the volume index of the prostate in aged mouse by long-term intragastrical administration.
Conclusion
The inhibitory effects by S-DOX on cardiovascular system are significantly weaker than those by rac-DOX and R-DOX, while S-DOX inhibits the vesical micturition pressure to the same extent as rac-DOX. Results of the present study suggest that a decrease in vesical micturition pressure induced by S-DOX is related to the reduction in contractile responses to electric field stimulation via its blocking action on the postsynapticα1-adrenoceptors in the urinary bladder detrusor smooth muscle and prostate smooth muscle, and S-DOX has the same affinity forα1-adrenoceptors as R-DOX in the isolated rabbit urinary bladder detrusor smooth muscle and prostate smooth muscle.
S-DOX, R-DOX and rac-DOX inhibit theα1-adrenoceptor activity to the same extent in the isolated rabbit urethral strips. However,α1-adrenoceptor antagonists of rac-DOX and prazosin are not able to affect the neurogenic contractions induced by electric field stimulation in the urethral strips. Neurogenic contractile responses to electric field stimulation in the isolated rabbit urethral strips are motor neuron-evoked contractions with characteristics of highly sensitive to nicotine and suramine.
第一部分多沙唑嗪光学异构体对兔离体膀胱逼尿肌的作用和作用机制
本研究观察了S-DOX、R-DOX和rac-DOX对兔离体膀胱逼尿肌收缩反应的影响。利用兔离体背侧和腹侧膀胱逼尿肌标本,分析多沙唑嗪及其光学异构体对药物或神经刺激诱发兔离体膀胱逼尿肌收缩反应的影响并分析其作用机制。
1背侧、腹侧膀胱逼尿肌对卡巴胆碱和苯肾上腺素诱发收缩反应的比较卡巴胆碱(0.01~30μmol?L-1)使背侧和腹侧膀胱逼尿肌产生浓度依赖性收缩反应,卡巴胆碱的浓度-反应曲线在两种标本上无显著性差异(P>0.05);与卡巴胆碱不同,苯肾上腺素(0.1~300μmol?L-1)仅使背侧膀胱逼尿肌产生浓度依赖性收缩反应,对腹侧膀胱逼尿肌无收缩作用。
2 S-DOX、R-DOX及rac-DOX对苯肾上腺素诱发兔背侧膀胱逼尿肌收缩反应的影响
苯肾上腺素使背侧膀胱逼尿肌产生浓度依赖性收缩反应,溶剂对苯肾上腺素诱发的兔背侧膀胱逼尿肌收缩反应无影响(P>0.05)。S-DOX、R-DOX和rac-DOX拮抗苯肾上腺素诱发兔背侧膀胱逼尿肌收缩反应的pKB值分别为7.44±0.19、7.39±0.14和7.38±0.30,三者之间无显著性差异(P>0.05)。
3 S-DOX、R-DOX及rac-DOX对电场刺激诱发兔背侧、腹侧膀胱逼尿肌收缩反应的影响
电场刺激诱发兔背侧和腹侧膀胱逼尿肌产生稳定的收缩反应,该收缩反应被0.1μmol?L-1的河豚毒素(TTX)完全阻断。S-DOX、R-DOX和rac-DOX均显著抑制电场刺激诱发的背侧膀胱逼尿肌收缩反应(P<0.01),且三者的抑制作用强度无显著性差异(P>0.05);但是,它们对电场刺激诱发的兔腹侧膀胱逼尿肌收缩反应无影响(P>0.05)。
上述结果表明,S-DOX、R-DOX和rac-DOX通过阻断膀胱逼尿肌α1受体显著抑制电场刺激诱发的兔背侧膀胱逼尿肌收缩反应,三者的抑制作用强度无显著性差异。
第二部分多沙唑嗪光学异构体对兔离体前列腺平滑肌收缩反应的影响
本研究观察了S-DOX、R-DOX和rac-DOX对兔离体前列腺收缩反应的影响。利用兔离体前列腺标本,分析多沙唑嗪及其光学异构体对药物或神经刺激诱发兔离体前列腺收缩反应的影响并分析其作用机制。
1 S-DOX、R-DOX及rac-DOX对苯肾上腺素诱发兔离体前列腺收缩反应的影响
苯肾上腺素使前列腺产生浓度依赖性收缩反应,溶剂对苯肾上腺素诱发的兔离体前列腺收缩反应无影响(P>0.05)。0.01μmol·L~(-1)和0.03μmol·L~(-1)的S-DOX、R-DOX或rac-DOX均可使苯肾上腺素量效曲线平行右移,Emax值不变。Schild plot分析结果表明,S-DOX、R-DOX或rac-DOX竞争性拮抗苯肾上腺素诱发的兔离体前列腺收缩反应,其pA2值分别为8.22±0.03、8.25±0.03和8.20±0.04,三者间无显著性差异(P>0.05)。S-DOX、R-DOX和rac-DOX拮抗苯肾上腺素诱发兔前列腺收缩反应的作用强度相同。
2 S-DOX、R-DOX及rac-DOX对电场刺激诱发兔离体前列腺收缩反应的影响
电场刺激诱发兔离体前列腺产生稳定的收缩反应,该收缩反应被0.1μmol·L~(-1)的TTX完全阻断。0.01μmol·L~(-1)或0.1μmol·L~(-1)的S-DOX、R-DOX和rac-DOX显著抑制电场刺激诱发的兔前列腺收缩反应(P<0.01),三者对电场刺激诱发兔离体前列腺收缩反应的抑制强度无显著性差异(P>0.05)。
3 Atr、Phent以及PPADS对电场刺激诱发兔离体前列腺收缩反应的影响
1μmol·L~(-1)Atr单独应用显著抑制电场刺激诱发的前列腺收缩反应,抑制百分率为62.6±11.7%(P<0.01),联合应用10μmol·L~(-1)的Phent后,二者完全抑制电场刺激诱发的前列腺收缩反应。单独给予10μmol·L~(-1)Phent可显著抑制电场刺激诱发的前列腺收缩反应,抑制百分率为96.7±9.3%(P<0.01),继续给予1μmol·L~(-1)Atr后,二者完全抑制电场刺激诱发的前列腺收缩反应。10μmol·L~(-1)PPADS对电场刺激诱发的前列腺收缩反应无明显影响(P>0.05)。TTX(0.1μmol·L~(-1))完全阻断该条件下电场刺激诱发的收缩反应。
上述结果表明,多沙唑嗪及其光学异构体显著抑制电场刺激诱发的兔离体前列腺收缩反应,其作用机制与药物阻断突出后α1受体有关,三者对兔离体前列腺α1受体的亲和力相同。
第三部分多沙唑嗪光学异构体对兔离体尿道环肌收缩功能的影响
制备兔离体膀胱颈和尿道环肌标本,记录药物或神经刺激诱发的收缩反应。观察多沙唑嗪及其光学异构体对苯肾上腺素诱发兔离体膀胱颈和尿道环肌收缩反应的影响;分析药物对电场刺激诱发兔离体尿道环肌收缩反应的作用及其作用机制。
1 S-DOX、R-DOX及rac-DOX对苯肾上腺素诱发兔膀胱颈和尿道环肌收缩反应的影响
苯肾上腺素使膀胱颈和各部位尿道环肌产生浓度依赖性收缩反应,溶剂对苯肾上腺素诱发的兔膀胱颈和尿道环肌收缩反应无影响(P>0.05)。0.03μmol·L~(-1)的S-DOX、R-DOX或rac-DOX拮抗苯肾上腺素诱发的兔离体膀胱颈收缩反应的pKB值分别为7.88±0.23、7.92±0.11和7.87±0.10,三者之间无显著性差异(P>0.05)。同样,在上段尿道、中段尿道和下段尿道,S-DOX、R-DOX和rac-DOX拮抗苯肾上腺素诱发各段尿道环肌收缩反应的pKB值亦无显著性差异(P>0.05)。
2 rac-DOX对电场刺激诱发兔膀胱颈和尿道环肌收缩反应的影响
溶剂对各标本的频率-反应曲线无明显影响(P>0.05)。不同浓度的rac-DOX(0.01、0.1和1μmol·L~(-1))对电场刺激诱发的兔离体膀胱颈和各段尿道环肌收缩反应无影响(P>0.05)。TTX(0.1μmol·L~(-1))完全阻断该条件下电场刺激诱发的收缩反应。
3 Pra、Atr及Ind对电场刺激诱发兔膀胱颈和尿道环肌收缩反应的影响
溶剂对各标本的频率-反应曲线无明显影响(P>0.05),TTX(0.1μmol·L~(-1))完全阻断电场刺激诱发的收缩反应。单独给予1μmol·L~(-1)的Atr显著抑制电场刺激诱发的兔膀胱颈收缩反应(P<0.05)。1μmol·L~(-1)的Pra对电场刺激诱发的膀胱颈收缩反应无显著影响;联合应用1μmol·L~(-1)的Atr后,抑制作用与单独给予Atr无显著性差异;与Pra和Atr联合用药组比,合用1μmol·L~(-1)的Ind进一步抑制了电场刺激诱发的兔离体膀胱颈收缩反应(P<0.01)。
在上段、中段以及下段尿道,1μmol·L~(-1)的Pra、Atr和Ind单独及联合应用均对电场刺激诱发的收缩反应无显著影响(P>0.05)。
4 Atr、Phent、Tub、尼古丁、苏拉明及TTX对电场刺激诱发兔尿道环肌收缩反应的影响
电场刺激诱发兔尿道环肌产生稳定的收缩反应。Atr(0.1~3μmol·L~(-1))和Phent(1~30μmol·L~(-1))对电场刺激诱发的兔尿道环肌收缩反应无影响。0.01μmol·L~(-1)和0.03μmol·L~(-1)的Tub显著抑制电场刺激诱发的收缩反应,抑制百分率分别为11.4±6.2%(P<0.05)和52.1±14.8%(P<0.01)。10μmol·L~(-1)的尼古丁显著抑制电场刺激诱发的收缩反应,抑制百分率为53.7±14.9%(P<0.01)。10、30、和100μmol·L~(-1)的苏拉明显著抑制电场刺激诱发的收缩反应,抑制百分率分别为22.5±3.5%、33.8±5.3%和54.2±5.9%。此外,0.1μmol·L~(-1)的Tub、30μmol·L~(-1)的尼古丁以及0.1μmol·L~(-1)的TTX尚能完全阻断电场刺激诱发的收缩反应。
5 CCh、Mox、Clo、Ado、ATP、UTP、5-HT和His对兔尿道环肌的作用
Mox(10μmol·L~(-1))和Clo(10μmol·L~(-1))可诱发尿道环肌产生较小收缩反应,标本最大收缩反应分别为0.20±0.12g和0.11±0.05g。CCh(30μmol·L~(-1))、Ado(30μmol·L~(-1))、ATP(10μmol·L~(-1))、UTP(30μmol·L~(-1))、5-HT(3μmol·L~(-1))和His(30μmol·L~(-1))对尿道标本均无收缩作用。
上述结果表明,S-DOX、R-DOX和rac-DOX通过阻断α1受体拮抗苯肾上腺素诱发的兔膀胱颈和尿道环肌收缩反应,三者阻断α1受体的作用相同。电场刺激诱发的兔尿道环肌收缩反应具有骨骼肌运动神经兴奋的特性,这一收缩成分同时易被尼古丁或苏拉明阻断。
第四部分多沙唑嗪光学异构体对老年小鼠前列腺增生的影响
实验采用老年小鼠长期(12周)灌胃给药法,观察了S-DOX、R-DOX和rac-DOX对老年小鼠前列腺体积、湿重、细胞增殖周期与凋亡百分率的影响。
1 S-DOX、R-DOX和rac-DOX对前列腺体积和湿重的影响与青年对照组相比,老年组小鼠前列腺体积指数无显著性差异,而S-DOX 6.0mg·kg-1组、R-DOX 6.0mg·kg-1组和R-DOX 0.6mg·kg-1组以及rac-DOX 6.0mg·kg-1组小鼠前列腺体指数显著减小(P<0.05)。
2 S-DOX、R-DOX和rac-DOX对小鼠前叶前列腺细胞增殖周期及凋亡的影响
与青年对照组相比,老年组小鼠前叶前列腺G0/G1期细胞周期百分率显著降低(P<0.01), G2/M期细胞周期百分率和增殖指数均显著增高(P<0.01);老年组小鼠前列腺细胞细胞凋亡率无明显改变。与老年组相比,S-DOX、R-DOX和rac-DOX老年小鼠前列腺细胞的增值指数和细胞周期无显著影响。
上述结果表明,长期(12周)灌胃给予S-DOX、R-DOX和rac-DOX,可能轻度降低老年小鼠前列腺细胞体积指数。
结论
S-DOX对心血管系统的抑制作用明显弱于R-DOX和rac-DOX,但是其降低膀胱排尿压的作用与同剂量rac-DOX相同。本研究证明,S-DOX降低动物膀胱排尿压的作用与其抑制去甲肾上腺素能神经兴奋所致的膀胱逼尿肌和前列腺平滑肌收缩反应有关;其作用机制是阻断了上述平滑肌组织的α1受体。S-DOX的上述作用及作用强度与R-DOX相同,说明兔膀胱逼尿肌和前列腺平滑肌细胞膜上的α1受体对多沙唑嗪光学异构体的亲和力相同。
S-DOX、R-DOX及rac-DOX可阻断兔尿道环肌α1受体,三者的作用强度相同;但是α1受体阻断药rac-DOX及哌唑嗪不影响电场刺激诱发的兔尿道环肌收缩反应。电场刺激诱发的兔尿道环肌收缩反应具有骨骼肌运动神经兴奋的特性,这一收缩成分同时易被尼古丁或苏拉明阻断。
Benign prostatic hyperplasia (BPH) often leads to lower urinary tract symptoms (LUTS). Patients with BPH/LUTS are suffering serious troubles on the quality of life. racemic-Doxazosin (rac-DOX),a long acting selectiveα1-adrenoceptor antagonist, is the current medication for the clinical treatment of BPH/LUTS. Several clinical studies have demonstrated that rac-DOX produces a decrease in urethral resistance and improve in lower urinary tract symptoms by blockingα1-adrenoceptor. However, rac-DOX is limited in the clinical application for BPH/LUTS due to its common adverse effects of dizziness, headache, and orthostatic hypotension. Therefore, the urinary tract selectivity should be necessarily considered whenα1-adrenoceptor antagonists are used for the BPH/LUTS patients.
rac-DOX is a mixture of S-doxazosin (S-DOX) and R-doxazosin (R-DOX). In a pharmacological study using the human isolated prostate tissue, S-DOX, R-DOX and rac-DOX all competitively antagonized the phenylephrine-induced contractile responses without significant differences in the pA2 values among S-DOX, R-DOX and rac-DOX. Recently, a study in our laboratory showed that the effects of S-DOX on animal blood pressure and onα1-adrenoceptors of the isolated rabbit blood vessels were significantly weaker than rac-DOX and R-DOX. However,the inhibitory effect by S-DOX on vesical micturition pressure was same to rac-DOX. To investigate the mechanisms of decreasing vesical micturition pressure by doxazosin and its enantiomers in animals, we observed the effects of S-DOX, R-DOX and rac-DOX on the contractile responses to drugs and electric field stimulation in the isolated rabbit urinary bladder detrusor, prostate and urethral strips, and the effects of S-DOX, R-DOX and rac-DOX on the prostate cell proliferation and apoptosis in the aged mouse.
PartⅠEffects of doxazosin and its enantiomers on the contractile responses in the isolated rabbit urinary detrusor strips
Contractile responses to drugs or electric field stimulation in the dorsal and ventral detrusor strips of the isolated rabbit urinary bladder were recorded to study the effects of S-DOX, R-DOX and rac-DOX on the contractile responses.
1 Contractile responses to carbachol and phenylephrine in the dorsal and ventral detrusor strips of the isolated rabbit urinary bladder
Carbachol produced contractile responses concentration-dependently in the dorsal and ventral detrusor strips, and the concentration-response curves for carbachol in the two kinds of strips were not significantly different from each other. Phenylephrine induced contractile responses in a concentration-dependent manner in the dorsal detrusor strips but not in the ventral detrusor strips.
2 Effects of doxazosin and its enantiomers on the contractile responses to phenylephrine in the dorsal detrusor strips of the isolated rabbit urinary bladder
Phenylephrine produced contractile responses concentration-dependently in the dorsal detrusor strips, and solvent had no effect on the contractile responses. The pKB values of S-DOX, R-DOX and rac-DOX at 1μmol·L~(-1) against phenylephrine in the dorsal detrusor strips of the isolated rabbit urinary bladder were 7.44±0.19, 7.39±0.14 and 7.38±0.30, respectively, and the pKB values of doxazosin and its enantiomers were not significantly different from each other (P>0.05).
3 Effects of doxazosin and its enantiomers on the contractile responses to electric field stimulation in the dorsal and ventral detrusor strips of the isolated rabbit urinary bladder
Electric field stimulation produced a steady contractile responses those were completely inhibited by tetrodotoxin (TTX) at 0.1μmol·L~(-1). S-DOX, R-DOX and rac-DOX significantly inhibited the contractile responses to electric field stimulation in the dorsal detrusor strips of the isolated rabbit urinary bladder (P<0.05), and their inhibitory effects were not significantly different from each other (P>0.05). However, S-DOX, R-DOX and rac-DOX did not affect the responses to electric field stimulation in the ventral detrusor strips.
These results indicate that S-DOX, R-DOX and rac-DOX significantly inhibit the contractile responses to electric field stimulation in the dorsal detrusor strips of the isolated rabbit urinary bladder via postsynapticα?1-adrenoceptors, and S-DOX inhibits the responses to the same extent as R-DOX and rac-DOX.
PartⅡEffects of doxazosin and its enantiomers on contractile responses in the isolated rabbit prostate strips
Contractile responses to drugs or electric field stimulation were recorded to study the effects of S-DOX, R-DOX and rac-DOX on the contractile responses in the isolated rabbit prostate strips.
1 Effects of doxazosin and its enantiomers on the contractile responses to phenylephrine in the isolated rabbit prostate strips
Phenylephrine produced contractile responses concentration-dependently in the prostate strips, and solvent did not affect the contractile responses. S-DOX, R-DOX and rac-DOX at 0.01μmol·L~(-1) and 0.03μmol·L~(-1) produced parallel shift of the cumulative concentration-response curves (CCRCs) for phenylephrine to the right in the isolated rabbit prostate, and the values of Emax of CCRCs for phenylephrine were not changed significantly by S-DOX, R-DOX and rac-DOX (P>0.05). The Schild plot analysis indicated that S-DOX, R-DOX and rac-DOX competitively antagonized the phenylephrine-induced contractile responses in the isolated rabbit prostate strips. The pA2 values of S-DOX, R-DOX and rac-DOX were 8.22±0.03, 8.25±0.03 and 8.20±0.04, respectively, and the pA2 values of three agents were not significantly different from each other (P>0.05).
2 Effects of doxazosin and its enantiomers on the contractile responses to electric field stimulation in the isolated rabbit prostate strips
Electric field stimulation produced a steady contractile responses, and the responses were completely inhibited by TTX at 0.1μmol·L~(-1). S-DOX, R-DOX and rac-DOX significantly inhibited the contractile responses to electric field stimulation in the isolated rabbit prostate strips (P<0.01), and S-DOX inhibited the responses to the same extent as R-DOX and rac-DOX.
3 Effects of atropine, phentolamine and PPADS on the contractile responses to electric field stimulation in the isolated rabbit prostate strips
Atropine (1μmol·L~(-1)) significantly reduced the contractile responses to electric field stimulation by 62.6±11.7%, and co-application with 10μmol·L~(-1) phetolamine abolished the contractile responses to electric field stimulation. Phetolamine (10μmol·L~(-1)) alone markedly reduced the contractile responses to electric field stimulation by 96.7±9.3%, and co-application with 1μmol·L~(-1) atropine abolished the contractile responses to electric field stimulation. PPADS (10μmol·L~(-1)) failed to affect the contraction induced by electric field stimulation in the isolated rabbit prostate strips (P>0.05). TTX (0.1μmol·L~(-1)) abolished the contractile responses.
These results indicate that doxazosin and its enantiomers significantly inhibit the contractile responses to electric field stimulation via postsynapticα1-adrenoceptors in the isolated rabbit prostate strips. The pA2 values of doxazosin and its enantiomers against phenylephrine are not significantly different from each other in the isolated rabbit prostate strips.
PartⅢEffects of doxazosin and its enantiomers on the contractile responses in the isolated rabbit urethral strips
In the urinary bladder neck strip and proximal, medial and distal urethral strips of the rabbit, the contractile responses to drugs and electric field stimulation were recorded. The effects of S-DOX, R-DOX and rac-DOX on the contractile responses to phenylephrine in the urinary bladdr neck strip and proximal, medial and distal urethral strips were observed. The pharmacological prolife of contractile responses to electric field stimulation in the urethral strips were investigated.
1 Effects of doxazosin and its enantiomers on the contractile responses to phenylephrine in the isolated rabbit urinary bladder neck strip and urethral strips
Phenylephrine produced contractile responses concentration-dependently in the isolated rabbit urinary bladder neck strip and the urethral strips, and solvent did not affect the contractile responses (P>0.05). The pKB values of S-DOX, R-DOX and rac-DOX against phenylephrine in the bladder neck were 7.88±0.23, 7.92±0.11 and 7.87±0.10, respectively, and the pKB values of the three agents were not significantly different from each other (P>0.05). The pKB values of doxazosin and its enantiomers against phenylephrine were not significantly different in the proximal, medial and distal urethral strips.
2 Effects of rac-DOX on the contractile responses to electric field stimulation in the isolated rabbit urinary bladder neck strip and proximal, medial and distal urethral strips
Solvent did not affect the frequency-response curves for electric field stimulation in the isolated rabbit urinary bladder neck strip and urethral strips. rac-DOX (0.01-1μmol·L~(-1)) was ineffective against the contractions induced by electric field stimulation in the isolated urinary bladder neck strip and proximal, medial and distal urethral strips, even though TTX at 0.1μmol·L~(-1) completely inhibited the responses.
3 Effects of prazosin, atropine and indomethacin on the contractile responses to electric field stimulation in the isolated rabbit urinary bladder neck strip and proximal, medial and distal urethral strips
Solvent did not affect the frequency-response curves for electric field stimulation in the isolated rabbit urinary bladder neck strip and urethral strips. Prazosin (1μmol·L~(-1)) was ineffective against the contractile responses to electric field stimulation (P>0.05), while atropine (1μmol·L~(-1)) significantly reduced the contractions (P<0.01). Co-application of prazosin and atropine caused an inhibition to the same extent as atropine alone (P>0.05). A treatment with the combination of prazosin, atropine and indomethacin decreased the contractile responses more potently than a treatment with a combination of prazosin and atropine (P<0.01). In the proximal, medial and distal urethral strips of the rabbit, prazosin, atropine or indomethacin did not affect the contractile responses to electric field stimulation, even though TTX at 0.1μmol·L~(-1) abolished the responses.
4 Effects of atropine, phentolamine, tubocurarine, nicotine, suramin and tetrodotoxin on the contractile responses to electric field stimulation in the isolated rabbit urethral strips
Electric field stimulation produced a steady contractile response in the isolated rabbit urethral strips. Atropine (0.1-3μmol·L~(-1)) or phentolamine (1-30μmol·L~(-1)) did not affect the contraction induced by electric field stimulation in the isolated rabbit urethral strips. Tubocurarine significantly inhibited the contraction at 0.01μmol·L~(-1) and 0.03μmol·L~(-1) by 11.4±6.2% and 52.1±14.8%, respectively. Nicotine at 10μmol·L~(-1) significantly inhibited the contraction by 53.7±14.9%. Suramin at 10, 30 and 100μmol·L~(-1) significantly inhibited the contraction by 22.5±3.5%, 33.8±5.3% and 54.2±5.9%, respectively.
Tubocurarine (0.1μmol·L~(-1)), Nicotine (30μmol·L~(-1)) and TTX (0.1μmol·L~(-1)) abolished the contractile responses to electric field stimulation in the isolated rabbit urethral strips. 5 Responses of the isolated rabbit urethral strips to carbachol, moxonidine, clonidine, adenosine, ATP, UTP, 5-HT and histamine
Moxonidine (10μmol·L~(-1)) and clonidine (10μmol·L~(-1)) produced small contractions in the isolated rabbit urethral strips, and the contractile force was 0.20±0.12g and 0.11±0.05g, respectively. Carbachol (30μmol·L-1), adenosine (30μmol·L~(-1)), ATP (10μmol·L~(-1)), UTP (30μmol·L~(-1)), 5-HT (3μmol·L~(-1)) and histamine (30μmol·L~(-1)) could not produce any contractile responses in the urethral strips.
These results indicate that doxazosin and its enantiomers antagonize the phenylephrine-induced contractile responses in the isolated rabbit urinary bladder neck strips and proximal, medial and distal urethral strips via postsynapticα1-adrenoceptors, and the pKB values of doxazosin and its enantiomers are not significantly different from each other.
Neurogenic contractile responses to electric field stimulation are motor neuron-evoked contractions with characteristics of highly sensitive to nicotine and suramine, and the neurogenic contractions are not affected by rac-DOX and prazosin.
PartⅣEffects of doxazosin and its enantiomers on the volume and cell proliferation of the prostate in aged mouse
The male mouse at 13 months of age was used to study the effects of long-term (12 weeks) intragastrical administration of doxazosin and its enantiomers on the volume and wet weight of the prostate, and the effects on the cell proliferation and apoptosis of the anterior lobe prostatic cells in aged mouse.
1 Effects of doxazosin and its enantiomers on the volume and wet weight of the prostate in aged mouse
In comparison with young mouse, the volume index of the prostate in aged mouse did not change significantly (P>0.05), but the volume index of the prostate in aged mouse treated with S-DOX (6.0mg·kg-1), R-DOX (6.0mg·kg-1), R-DOX (0.6mg·kg-1) or rac-DOX (6.0mg·kg-1) was significantly decreased (P<0.05).
2 Effects of doxazosin and its enantiomers on the cell cycle, proliferation and apoptosis of the anterior lobe cells of the prostate in aged mouse
In comparison with young mouse, the total G0/G1-phase of the cells of the anterior lobe of the prostate were significantly decreased (P<0.01), and the total G2/M-phase cells of the anterior lobe of the prostate and the proliferation index of the prostatic cells were significantly increased in aged mouse (P<0.01).
In comparison with aged mouse, the proliferation index and cell cycle distribution of the anterior prostatic lobe in aged mouse treated by S-DOX, R-DOX and rac-DOX did not change significantly (P>0.05).
These results indicate that doxazosin and its enantiomers slightly decrease the volume index of the prostate in aged mouse by long-term intragastrical administration.
Conclusion
The inhibitory effects by S-DOX on cardiovascular system are significantly weaker than those by rac-DOX and R-DOX, while S-DOX inhibits the vesical micturition pressure to the same extent as rac-DOX. Results of the present study suggest that a decrease in vesical micturition pressure induced by S-DOX is related to the reduction in contractile responses to electric field stimulation via its blocking action on the postsynapticα1-adrenoceptors in the urinary bladder detrusor smooth muscle and prostate smooth muscle, and S-DOX has the same affinity forα1-adrenoceptors as R-DOX in the isolated rabbit urinary bladder detrusor smooth muscle and prostate smooth muscle.
S-DOX, R-DOX and rac-DOX inhibit theα1-adrenoceptor activity to the same extent in the isolated rabbit urethral strips. However,α1-adrenoceptor antagonists of rac-DOX and prazosin are not able to affect the neurogenic contractions induced by electric field stimulation in the urethral strips. Neurogenic contractile responses to electric field stimulation in the isolated rabbit urethral strips are motor neuron-evoked contractions with characteristics of highly sensitive to nicotine and suramine.
引文
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2 Morgan CW, de Groat WC, Felkins LA, et al. Intracellular injection of neurobiotin or horseradish peroxidase reveals separate types of preganglionic neurons in the sacral parasympathetic nucleus of the cat. J Comp Neurol, 1993, 331:161~182
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