Human chorionic gonadotrophin (hCG) secretion by the syncytiotrophoblast of the human placenta is essential for a successful pregnancy. It has previously been demonstrated that extracellular Ca²⁺ is necessary for hCG secretion (Sharma & Rao, 1997) and entry of Ca²⁺ via L-type Ca²⁺ channels has been implicated in this process (Sharma & Rao, 1997; Cemerik et al. 1998). However, direct studies have failed to identify L-type Ca²⁺ channels in human placental trophoblast cells (Bax et al. 1994; Cronier et al. 1999). Therefore the entry pathway for Ca²⁺ is not clear. The aim of this study was to examine the effect of a range of Ca²⁺-permeable channel blockers on hCG secretion from human placental villous fragments.

With ethical approval, villous fragments of approximately 3-5 mg wet weight were dissected from term human placentas. Fragments were washed in control Tyrode solution (mM: NaCl 135, KCl 5, MgCl₂ 1, CaCl₂ 1.8, glucose 5.6, Hepes 10; pH 7.4 with NaOH), then incubated (six per vial) for 1 h in Ca²⁺-free (0 mM CaCl₂ + 0.5 mM EGTA), control Tyrode buffer or buffer containing 10 mM CaCl₂ in the presence and absence of three Ca²⁺-permeable channel blockers. The blockers examined were: 50 µM SKF96365 (blocks store-operated Ca²⁺ channels), 150 µM GdCl₃ (a blocker of Ca²⁺ entry pathways including non-selective cation channels), 1 µM nifedipine (blocks L-type Ca²⁺ channels). Following incubation hCG secretion was assessed by assay and placental fragment blotted wet weight was determined. Data were calculated as mIU ml^-1 mg^-1 wet weight, then normalised to the control hCG secretion to account for interplacental variability.

hCG secretion in Ca²⁺-free buffer was significantly reduced compared with control, whereas there was a significant increase in hCG secretion following exposure to 10 mM CaCl₂ (Fig. 1). hCG secretion stimulated by 10 mM CaCl₂ was significantly reduced in the presence of 150 µM GdCl₃; nifedipine and SKF6936 were without effect (see Fig. 1).

We have confirmed that hCG secretion by human term placenta is dependent on extracellular Ca²⁺. The data suggest that L-type Ca²⁺ channels are not involved in regulation of hCG secretion. Conversely the effect of GdCl₃ suggests that extracellular Ca²⁺ entry via Ca²⁺-permeable non-selective cation channels may be important in this process.

This work was supported by the MRC.

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Figure 1. Graph showing normalised data for hCG secretion from term placental villous fragments. Values are medians ± range, n = 6. *P < 0.05 vs. control; #P < 0.05 vs. 10 mM Ca2+ (Kruskal-Wallis with Dunn’s multicomparisons test). con, control; 0Ca, Ca2+-free buffer; 10Ca, 10 mM Ca2+ buffer; SKF, 50 µM SKF96365; Gd, 150 µM GdCl3; nif, 1 µM nifedipine (all channel blockers were in 10 mM Ca2+ buffer).