Robberecht W, Andries M, Denef C
Laboratory of Cell Pharmacology, University of Leuven, School of Medicine, Belgium.
Neuroendocrinology. 1992 Oct;56(4):550-60. doi: 10.1159/000126273.
Angiotensin II (AII)-like immunoreactivity (LIR) was detected by immunostaining in 7.5 +/- 1.1% of cells obtained by redispersion of pituitary cell aggregates from 15- to 20-day-old female rats, cultured for 5-7 days in serum-free medium supplemented with thyroid hormone and dexamethasone. Also, renin-LIR was retained in these cultures. As shown by double immunostaining of paraffin-embedded sections of the aggregates, this AII-LIR was localized only in gonadotrophs. AII-LIR was detected at least up to 5 weeks in culture. On reversed-phase, high-performance liquid chromatography (HPLC), this AII-LIR co-migrated with authentic AII. In perifused aggregate cell cultures of 15- to 20-day-old female rat pituitary maintained in serum-free medium supplemented with dexamethasone (DEX) and triiodothyronine (T3), AII stimulated GH release. AI and AIII had a similar effect. To evaluate the possible involvement of endogenous AII in the local regulation of GH release, gonadotrophs were stimulated with luteinizing hormone-releasing hormone (LHRH). LHRH displayed a transient inhibitory effect on GH release, which was followed by a rebound of GH release after withdrawal of the peptide. Treatment of aggregates with pertussis toxin reversed this inhibitory effect into a significant stimulation of GH release. In aggregates cultured in serum-supplemented medium, LHRH provoked a significant stimulation of GH release which was still followed by a post-stimulus rebound release. In hemipituitaries from 5-day-old rats, a significant stimulatory effect of LHRH on GH release was found without rebound secretion. To evaluate the possible involvement of endogenous AII in the effects of LHRH on GH release, the influence of (Sar1,Ala8)AII, a peptide AII receptor antagonist, and of DUP753, a non-peptide AII receptor blocker was tested in various in vitro conditions. The effect of LHRH on GH release in aggregates cultured either in serum-free medium supplemented with DEX and T3 or in serum-supplemented medium was not affected by (Sar1,Ala8)AII, not even after enhancing the LHRH-induced GH release by treatment of the aggregates with pertussis toxin. A hundred times lower concentration of (Sar1,Ala8)AII, however, abolished the AII-induced changes in GH release. Also DUP753 (10 microM) failed to block LHRH-induced GH release in aggregates. (Sar1,Ala8)AII also failed to block the effect of LHRH on GH release from hemipituitaries. It is concluded that LHRH has inhibitory and stimulatory effects on GH release in cultured pituitary cell aggregates.(ABSTRACT TRUNCATED AT 400 WORDS)
通过免疫染色检测到，在15至20日龄雌性大鼠垂体细胞聚集体经再分散后获得的细胞中，7.5±1.1%的细胞有血管紧张素II（AII）样免疫反应性（LIR），这些细胞在补充了甲状腺激素和地塞米松的无血清培养基中培养5至7天。此外，这些培养物中保留了肾素-LIR。如聚集体石蜡包埋切片的双重免疫染色所示，这种AII-LIR仅定位于促性腺激素细胞中。在培养中至少5周都能检测到AII-LIR。在反相高效液相色谱（HPLC）上，这种AII-LIR与 authentic AII共迁移。在补充了地塞米松（DEX）和三碘甲状腺原氨酸（T3）的无血清培养基中维持培养的15至20日龄雌性大鼠垂体的灌流聚集体细胞培养物中，AII刺激生长激素（GH）释放。AI和AIII有类似作用。为评估内源性AII可能参与GH释放的局部调节，用促黄体生成素释放激素（LHRH）刺激促性腺激素细胞。LHRH对GH释放表现出短暂抑制作用，在肽撤出后随后是GH释放的反弹。用百日咳毒素处理聚集体将这种抑制作用转变为对GH释放的显著刺激。在补充血清的培养基中培养的聚集体中，LHRH引起GH释放的显著刺激，随后仍然是刺激后反弹释放。在5日龄大鼠的半垂体中，发现LHRH对GH释放有显著刺激作用且无反弹分泌。为评估内源性AII可能参与LHRH对GH释放的作用，在各种体外条件下测试了肽AII受体拮抗剂（Sar1，Ala8）AII和非肽AII受体阻滞剂DUP753的影响。在补充了DEX和T3的无血清培养基或补充血清的培养基中培养的聚集体中，LHRH对GH释放的作用不受（Sar1，Ala8）AII影响，即使在用百日咳毒素处理聚集体增强LHRH诱导的GH释放后也是如此。然而，低100倍浓度的（Sar1，Ala8）AII消除了AII诱导的GH释放变化。同样，DUP753（10 microM）未能阻断聚集体中LHRH诱导的GH释放。（Sar1，Ala8）AII也未能阻断LHRH对半垂体GH释放的作用。得出结论，LHRH对培养的垂体细胞聚集体中的GH释放有抑制和刺激作用。（摘要截断于400字）
