Poulin B, Rich N, Mas J L, Kordon C, Enjalbert A, Drouva S V
Unité de Dynamique des Systèmes Neuroendocriniens, U159 INSERM, Centre Paul Broca, Paris, France.
Mol Cell Endocrinol. 1998 Jul 25;142(1-2):99-117. doi: 10.1016/s0303-7207(98)00114-2.
Exposure of the gonadotrope cells to gonadotropin-releasing hormone (GnRH) reduces their responsiveness to a new GnRH stimulation (homologous desensitization). The time frame as well as the mechanisms underlying this phenomenon are yet unclear. We studied in a gonadotrope cell line (alphaT3-1) the effects of short as well as long term GnRH pretreatments on the GnRH-induced phospholipases-C (PLC), -A2 (PLA2) and -D (PLD) activities, by measuring the production of IP3, total inositol phosphates (IPs), arachidonic acid (AA) and phosphatidylethanol (PEt) respectively. We demonstrated that although rapid desensitization of GnRH-induced IP3 formation did not occur in these cells, persistent stimulation of cells with GnRH or its analogue resulted in a time-dependent attenuation of GnRH-elicited IPs formation. GnRH-induced IPs desensitization was potentiated after direct activation of PKC by the phorbol ester TPA, suggesting the involvement of distinct mechanisms in the uncoupling exerted by either GnRH or TPA on GnRH-stimulated PI hydrolysis. The levels of individual phosphoinositides remained unchanged under any desensitization condition applied. Interestingly, while the GnRH-induced PLA2 activity was rapidly desensitized (2.5 min) after GnRH pretreatments, the neuropeptide-evoked PLD activation was affected at later times, indicating an important time-dependent contribution of these enzymatic activities in the sequential events underlying the GnRH-induced homologous desensitization processes in the gonadotropes. Under GnRH desensitization conditions, TPA was still able to induce PLD activation and to further potentiate the GnRH-evoked PLD activity. AlphaT3-1 cells possess several PKC isoforms which, except PKCzeta, were differentially down-regulated by TPA (PKCalpha, betaII, delta, epsilon, eta) or GnRH (PKCbetaII, delta, epsilon, eta). In spite of the presence of PKC inhibitors or down-regulation of PKC isoforms by TPA, the desensitizing effect of the neuropeptide on GnRH-induced IPs, AA and PEt formation remained unchanged. In conclusion, in alphaT3-1 cells the GnRH-induced homologous desensitization affects the GnRH coupling with PLC, PLA2 and PLD by mechanism(s) which do not implicate TPA-sensitive PKC isoforms, but likely reflect time-dependent modification(s) on the activation processes of the enzymes.
促性腺激素细胞暴露于促性腺激素释放激素（GnRH）会降低其对新的GnRH刺激的反应性（同源脱敏）。这种现象的时间框架以及潜在机制尚不清楚。我们在一种促性腺激素细胞系（αT3-1）中研究了短期和长期GnRH预处理对GnRH诱导的磷脂酶-C（PLC）、-A2（PLA2）和-D（PLD）活性的影响，分别通过测量肌醇三磷酸（IP3）、总肌醇磷酸（IPs）、花生四烯酸（AA）和磷脂酰乙醇（PEt）的产生来进行。我们证明，尽管在这些细胞中未发生GnRH诱导的IP3形成的快速脱敏，但用GnRH或其类似物持续刺激细胞会导致GnRH引发的IPs形成随时间衰减。佛波酯TPA直接激活蛋白激酶C（PKC）后，GnRH诱导的IPs脱敏增强，这表明GnRH或TPA对GnRH刺激的磷脂酰肌醇水解的解偶联作用涉及不同机制。在任何应用的脱敏条件下，单个磷酸肌醇的水平均保持不变。有趣的是，虽然GnRH预处理后GnRH诱导的PLA2活性迅速脱敏（2.5分钟），但神经肽诱发的PLD激活在较晚时间受到影响，这表明这些酶活性在促性腺激素细胞中GnRH诱导的同源脱敏过程的连续事件中具有重要的时间依赖性作用。在GnRH脱敏条件下，TPA仍能够诱导PLD激活并进一步增强GnRH诱发的PLD活性。αT3-1细胞具有几种PKC同工型，除PKCζ外，它们被TPA（PKCα、βII、δ、ε、η）或GnRH（PKCβII、δ、ε、η）差异性下调。尽管存在PKC抑制剂或TPA对PKC同工型的下调，但神经肽对GnRH诱导的IPs、AA和PEt形成的脱敏作用保持不变。总之，在αT3-1细胞中，GnRH诱导的同源脱敏通过不涉及TPA敏感的PKC同工型的机制影响GnRH与PLC、PLA2和PLD的偶联，但可能反映了对酶激活过程的时间依赖性修饰。
