Chauhan M, Gangula P R R, Wimalawansa S J, Yallampalli C
Department of Obstetrics and Gynecology, University of Texas Medical Branch, Galveston, Texas 77555-1062, USA.
Biol Reprod. 2004 Jun;70(6):1658-63. doi: 10.1095/biolreprod.103.023895. Epub 2004 Feb 11.
The vascular relaxation sensitivity to calcitonin gene-related peptide (CGRP) is enhanced during pregnancy, compared with nonpregnant human and rat uterine arteries. In the rat uterine artery, two types of CGRP receptors have been shown to coexist, CGRP-A receptor, which is a complex of calcitonin receptor-like receptor (CRLR), and receptor activity-modifying protein (RAMP(1)) and CGRP-B receptor, which is different from CRLR. In the present study, we hypothesized that: 1) CGRP-induced vasorelaxation in rat uterine artery is mediated through CGRP-A receptor and 2) N-terminal (Nt) domain of CRLR (Nt-CRLR) has a major contribution in ligand binding and mediating CGRP- induced relaxation effects in rat uterine artery. Polyclonal antibodies against Nt-domain of CRLR and RAMP(1) (Nt-RAMP(1)) were raised in rabbits and characterized for their specificity and were used to inhibit CGRP-induced vasorelaxation in rat uterine artery. For vascular relaxation studies, uterine arteries from Day 18 pregnant rats were isolated, and responsiveness of the vessels to CGRP was examined with a small vessel myograph. CGRP (10(-10) to 10(-7) M) produced a concentration-dependent relaxation of norepinephrine-induced contractions in Day 18 pregnant rat uterine arteries. These effects were significantly (P < 0.05) inhibited when uterine arteries were incubated with the antibody raised against Nt-CRLR (PD(2) = 6.75 +/- 0.20) and were totally abolished in presence of antibodies for both Nt-CRLR and Nt-RAMP(1) (PD(2) = 6.14 +/- 0.35). In contrast, a monoclonal antibody for CGRP-B receptor had no effect on CGRP-induced rat uterine artery relaxation. These studies suggest that CGRP effects in rat uterine artery are mediated through CGRP-A receptor and that Nt-domain of CRLR may play a predominant role in CGRP binding and thus in causing CGRP-induced uterine artery relaxation.
与未怀孕的人和大鼠子宫动脉相比，在怀孕期间，血管对降钙素基因相关肽（CGRP）的舒张敏感性增强。在大鼠子宫动脉中，已显示两种类型的CGRP受体共存，即CGRP-A受体，它是降钙素受体样受体（CRLR）与受体活性修饰蛋白（RAMP(1)）的复合物；以及CGRP-B受体，它与CRLR不同。在本研究中，我们假设：1）CGRP诱导的大鼠子宫动脉血管舒张是通过CGRP-A受体介导的；2）CRLR的N末端（Nt）结构域在配体结合以及介导CGRP诱导的大鼠子宫动脉舒张效应中起主要作用。针对CRLR的Nt结构域和RAMP(1)（Nt-RAMP(1)）的多克隆抗体在兔中制备，并对其特异性进行了表征，用于抑制CGRP诱导的大鼠子宫动脉血管舒张。对于血管舒张研究，分离出妊娠第18天大鼠的子宫动脉，并用小型血管肌动描记器检测血管对CGRP的反应性。CGRP（10(-10)至10(-7) M）使妊娠第18天大鼠子宫动脉中去甲肾上腺素诱导的收缩产生浓度依赖性舒张。当子宫动脉与针对Nt-CRLR产生的抗体孵育时，这些效应被显著抑制（P < 0.05）（PD(2) = 6.75 +/- 0.20），并且在存在针对Nt-CRLR和Nt-RAMP(1)的抗体时完全消除（PD(2) = 6.14 +/- 0.35）。相比之下，针对CGRP-B受体的单克隆抗体对CGRP诱导的大鼠子宫动脉舒张没有影响。这些研究表明，CGRP在大鼠子宫动脉中的作用是通过CGRP-A受体介导的，并且CRLR的Nt结构域可能在CGRP结合以及由此导致CGRP诱导的子宫动脉舒张中起主要作用。
