Block of Recombinant T-Type Calcium Channels by Gossypol, a Potential Contraceptive

Gossypol is a bioactive compound associated with some male contraceptive properties. Experimental evidences have shown that gossypol inhibits spermatogenesis as well as spermatozoa motility and viability. T-type calcium channels, or CaV3 channels, have been found in both spermatogenic and sperm mammalian cells; they play an essential role in spermatogenesis, motility, capacitation and acrosome reaction. Recent studies show that native T-type calcium currents from murine spermatogenic cells are inhibited by micromolar concentrations of gossypol in a non-reversible process. However, until now there was no data showing whether gossypol preferentially blocks any member of the CaV3 channels subfamily, neither information about the blocking mechanism. Here, we performed whole-cell patch-clamp recordings of HEK-293 cells stably expressing human CaV3 channels (CaV3.1, CaV3.2 and CaV3.3) to address these questions. We found that gossypol blocks the three CaV3 channels in a concentration-response manner with similar IC50s ranging from 6 to 11 μM. Block by gossypol is not completely irreversible although T-type currents were poorly recovered (10-30%). Voltage-dependence of activation was not modified by gossypol, but steady-state inactivation was shifted to more negative potentials (around 10 mV), and block at −30 mV was stronger when using more depolarized holding potentials. In addition, the inactivated-state of CaV3 channels was stabilized in the presence of gossypol as the recovery from inactivation was delayed and incomplete. A modest use-dependent component was observed only in CaV3.1. Current kinetics were not affected by gossypol. The results demonstrate that gossypol blocks human CaV3 channels at micromolar concentrations with preferential binding to inactivated-states, and suggest that sperm cells will be more sensitive to gossypol during the hyperpolarization that take place along the capacitation process. These results provide new insights about the potential use of gossypol as a contraceptive.Supported by CONACYT-MEXICO 167790.