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In order to investigate the effect of leptin on the secretion of rat pituitary adenoma GH3 cell and its mechanisms, we observed the effect of leptin on the growth hormone secretion, proliferation and apoptosis of GH3 cells. The results indicated that leptin at 1, 10, and 100 nmol/L could inhibit the basal growth hormone secretion of GH3 cells in a dose dependent manner (P<0.05). Short-term treatment of leptin (10 nmol/L) for 30 min, 1 and 3 h did not affect basal GH secretion. However, treatment of the GH3 cells with leptin (10 nmol/L) for 1 d or longer resulted in an inhibition of GH secretion (P<0.05). We used MTT method and flow cytometery (FCM) to study the effect of leptin on the proliferation and apoptosis of GH3 cells. We found that leptin inhibited proliferation of GH3 cells with a dose-dependent manner. And leptin reduced the proportion of cells in S phase, increased the proportion of cells in G1, and increased the proportion of GH3 cells in 2 and 4 phase. These results demonstrate that leptin inhibits the basal GH secretion of GH3 cells, which may be due to the inhibition of DNA synthesis and advanced apoptosis of GH3 cells.

We found previously that ACh can significantly inhibit the proliferation of cultured human pituitary adenoma cells. In order to make a further investigation of the mechanism of the inhibitory effect of ACh on the proliferation of pituitary adenoma cells, we observed the levels of protein kinase C (PKC), [Ca(2+)](i) and cAMP/cGMP in cultured pituitary adenoma cells after treatment with ACh. The results demonstrate that (1) compared with control, PMA, a PKC activator, increased the activity of cytoplasm, membrane and total PKC in human pituitary adenoma cells. However, after a 15-min treatment with ACh (10 micromol/L), a significant reduction of the activity of cytoplasm, membrane and total PKC in human pituitary adenoma cells was observed, and the reduction effect could be blocked by atropine. (2) The level of [Ca(2+)](i) of single adenoma cells was found to decrease immediately on the addition of ACh (10 micromol/L), which could also be blocked by atropine. (3) ACh increased the amount of cAMP in the cytoplasm of human pituitary adenoma cells, but had no effect on that of cGMP. These data provide an important clue to explore the molecular mechanisms of the inhibitory effect of ACh on the proliferation of pituitary adenoma cells, and suggest that the modulating effect of ACh on the proliferation of pituitary adenoma cells results from the interactions of several cellular signaling pathways.

In order to elucidate the effect of acetylcholine (ACh) on the occurrence and development of human pituitary adenoma, it was firstly observed whether there exists choline acetyl transferase (ChAT) which is necessary for the synthesis of acetylcholine in the cells of human pituitary adenoma, and then MTT method, (3)H TdR incorporation, cell cycle analysis and TUNEL were employed to estimate the influence of ACh on the proliferation, DNA synthesis and apoptosis of three kinds of human pituitary adenoma (human prolactinoma, somatotropinoma and non-functional tumor) cells cultured in vitro. The results showed that (1) the positive staining of ChAT was obviously observed in the cells of the three kinds of human pituitary adenoma, however, it was lower than that in normal human pituitary gland; (2) ACh had a similar effect on the proliferation of the three kinds of human pituitary adenoma cells. ACh at 0.1-10 micromol/L decreased the (3)H TdR incorporation and the MTT A value in a dose-dependent manner. At the same time, ACh decreased the ratio of S or G(2) phase pituitary adenoma cells significantly, but increased the ratio of G(1) phase pituitary tumour cells markedly; (3) the effect of acetylcholine on the proliferation of human pituitary adenoma cells was inhibited by atropine, but not by tubocurarine; (4) ACh had no effect on the apoptosis of human pituitary adenoma cells cultured in vitro. These data suggest that ACh may have a significant modulating effect on the proliferation of pituitary adenoma cells by means of paracrine or autocrine, and the effect is mediated by muscarinic receptor.