Progesterone and CatSper dependency

Progesterone is present in high concentrations (μm) in the vicinity of the cumulus-oophorous complex and considered a key signalling factor for human spermatozoa. Progesterone induces an instant increase in intracellular calcium concentration [Ca²⁺]_i despite the absence of the classical progesterone receptor in human spermatozoa. Progesterone causes changes in beating of the flagellum but can also induce acrosome reaction in human spermatozoa. Both these effects are mediated by increased intracellular Ca²⁺ but they are exerted at opposite ends of this highly polarized cell. A breakthrough came from two recent studies (Lishko et al., 2011; Strunker et al., 2011) which showed that progesterone causes activation of the Ca²⁺-permeable CatSper channel in human spermatozoa, suggesting that CatSper or an associated protein serves as the long sought binding site for progesterone in human spermatozoa.

Progesterone not only raises [Ca²⁺]_i in human spermatozoa, it also activates a complex series of signalling events and cell activities. In this issue of the International Journal of Andrology, Sagare-Patil et al. (2012) report that some of these progesterone-mediated effects have characteristics that suggest they are activated by mechanisms independent of CatSper. Sagare-Patil and colleagues used a classical approach to investigate the concentration and time-dependent effects of progesterone. They report clear differences in dose dependence between different responses to progesterone. Motility stimulation, hyperactivation, tyrosine kinase activity, ERK1/2 phosphorylation and P90RSK phosphorylation all showed dose-dependence over 0.01–1 μm, consistent with the action of progesterone on CatSper. Acrosome reaction, tyrosine phosphorylation (analysed from immunofluorescence), P38MAPK phosphorylation, JNK phosphorylation and AKT phosphorylation were dose-dependently induced over the range 1–10 μm progesterone, indicating involvement of a lower affinity binding site for progesterone. These findings are in line with a previous report that progesterone binding and Ca²⁺ mobilization in human spermatozoa occur at nm progesterone (consistent with CatSper), but that a second low-affinity phase requires ≥5 μm progesterone (Luconi et al., 1998).

CatSper is the only identified calcium-permeable channel that has been detected by patch clamp of human spermatozoa (Kirichok & Lishko, 2011). T-type channel blockers NNC 55–0396 (2 μm) and mibefradil (30 μm) abolish CatSper currents in patch clamped human spermatozoa (Lishko et al., 2011; Strunker et al., 2011). These compounds also significantly inhibited the progesterone-induced Ca²⁺ response, but the signal was not abolished (Strunker et al., 2011). Sagare-Patil et al. (2012) also found that mibefradil (40 μm) was unable to abrogate the calcium increase induced by progesterone (∼80% reduction). In addition, mibefradil only partially inhibited the induction by progesterone of tyrosine phosphorylation and acrosome reaction. Likewise, tyrosine phosphorylation and the acrosome reaction are not diminished in CatSper knockout mouse spermatozoa and a late phase of the ZP-induced Ca²⁺ increase persists, although at lower frequency, in these cells (Ren & Xia, 2010). These data suggest that an as yet unidentified Ca²⁺-permeable channel exists in spermatozoa or that significant Ca²⁺ increases can originate from intracellular calcium stores. Ca²⁺ storage organelles are present at the sperm neck (Ho & Suarez, 2003; Costello et al., 2009) and thus a second site of progesterone-induced Ca²⁺ mobilization may occur here. Activated vitamin D (1,25(OH)₂D₃) appears to work by directly mobilizing stored Ca²⁺ at the sperm neck. This effect of 1,25(OH)₂D₃ on [Ca²⁺]_i is smaller than the effect of progesterone but has significant effects on sperm motility and acrosome reaction (Aquila et al.,2009; Blomberg Jensen et al., 2011). 1,25(OH)₂D₃ differs from progesterone and other activators of CatSper, as the response is mediated by the vitamin D receptor, depends solely on stored intracellular calcium and does not influence the responsiveness to progesterone (Blomberg Jensen et al., 2011). Whether progesterone can directly activate Ca²⁺ release here remains unclear. Kinetics of the [Ca²⁺]_i rise in response to progesterone are similar throughout the cell (Blomberg Jensen & Dissing, 2012; Blomberg Jensen et al., 2012; Servin-Vences et al., 2012), which is not consistent with a single site of Ca²⁺ influx in the flagellum. However, rapid buffering of extracellular Ca²⁺ by application of BAPTA simultaneously with progesterone (to avoid depletion of Ca²⁺ stores) completely abolished the progesterone-induced Ca²⁺ signal (Strunker et al., 2011).

In conclusion, the findings published by Sagare-Patil et al. (2012) emphasize that the action of progesterone on human spermatozoa is complex and suggest that some of the progesterone-activated effects may not be CatSper dependent. Thus, progesterone signalling may occur by two diverse mechanisms and sites of action: a high affinity binding through CatSper in the tail and possibly a second low-affinity binding site that may be elsewhere in the spermatozoa – the neck/head region being a possibility. There is also a possibility that some progesterone-induced events in human spermatozoa are fully calcium independent. We encourage future studies to focus on the spatial organization of signalling in human spermatozoa and to determine whether there exists another binding site/channel besides CatSper for progesterone, while we await development of specific inhibitors of CatSper.