Wataru Takahashi

Developmental changes in the functional, biochemical and molecular properties of rat bladder endothelin receptors

The effect of aging on functional, biochemical, anatomical and molecular properties of endothelin (ET) receptors in bladder smooth muscle of the 3-week-, 3-month- and 22-month-old rats was examined using isolated muscle bath techniques, radioligand binding on membrane particulates and slide mounted tissue sections, and real-time reverse transcription polymerase chain reaction (RT-PCR). ET-1 induced significantly larger contractile responses in bladder dome muscle strips from 3-week- than from 3-month- and 22-month-old rats. The expression level of total ET receptors, determined by saturation binding experiments with [125I]ET-1, was higher in detrusor from 3-week- than 22-month-old rats. Inhibition studies with BQ123, a selective ETA receptor antagonist, indicated the predominance of the ETA receptor subtype and a similar proportion of ETA to ETB receptor subtypes in the rat detrusor at all ages studied. Autoradiographic data support the age-dependent decrease in the density of ET receptors and also indicate that the ETA receptor subtype is primarily located in the smooth muscle layer, whereas the ETB receptor subtype is located in both the urothelial and smooth muscle layers. Determined by real-time RT-PCR, ET 1, ET-3, ECE-1 and ET receptor subtype (ETA and ETB) mRNAs were shown to be higher in bladders of 3-week- compared to 3-month- or 22-month-old rats. This study indicates age-dependent alterations in the ET receptor system at both gene transcript and protein levels in the Fischer rat detrusor.

The effect of castration on endothelins, their receptors and endothelin converting enzyme in rat prostate

Abstract. We previously have shown that experimental diabetes in rats causes prostatic involution, reduces serum testosterone levels, and causes an upregulation in prostatic endothelin (ET) receptors. Furthermore, insulin treatment normalizes these changes (Saito et al., Mol Cell Biochem 210:1–12, 2000). Since experimental diabetes-induced reduction in serum testosterone may be a factor in the alteration of the ET receptors and of prostatic growth, we investigated the effect of castration, another means of involuting the prostate and decreasing serum testosterone levels, on the expression of ET receptors in ventral and dorsolateral regions of the rat prostate.Three-month-old Sprague-Dawley rats were surgically castrated or sham operated, and then killed on the 7th post-operative day. Biochemical and pharmacological properties, and localization of ET receptors in the rat prostate, were determined by performing a series of binding experiments with [125I]ET-1 and by light microscopy autoradiography, respectively. The expression levels of ET-1, ET-3, ET receptor subtypes and endothelin converting enzyme-1 (ECE-1) mRNAs were assessed by relative multiplex reverse transcription polymerase chain reaction (RT-PCR).The total density of ET receptors increases 3.7-fold in the ventral and 2.1-fold in the dorsolateral regions of the castrated rat prostate compared to sham operated animals. Castration causes a 2.4-fold increase in the density of α1-adrenoceptors (α1-ARs) in the ventral region of the prostate, but no change in the density of α1-ARs in the dorsolateral region of the rat prostate. The predominant ET receptor subtype in the rat prostate is the ETA subtype, which is mainly located in the prostatic stroma. In addition, RT-PCR data show an upregulation in the expression of ETB receptor subtype, ET-1 and ECE-1 mRNA in both regions, and a downregulation in the expression of ETA receptor subtype mRNA in the dorsolateral region of the castrated rat prostate. There is no change in the expression of ET-3 mRNA in either region. Castration does not cause significant changes in the pharmacological properties of prostatic ET receptors, i.e., the predominance of ETA receptors in either region of the prostate, or the expression of ETA receptor subtype mRNA in the ventral region of the castrated rat prostate. These results suggest the existence of a region/lobe-specific regulatory role for testosterone in the expression of the ET receptor system in the rat prostate.

Regulatory effect of castration on endothelins, their receptors and endothelin‐converting enzyme in rat seminal vesicle

To investigate the effects of castration on the expression of endothelins (ETs), ET receptors and ET converting enzyme‐1 (ECE‐1) in the rat seminal vesicle (RSV).