The clinical significance of calcium-signalling pathways mediating human sperm hyperactivation

Abstract

STUDY QUESTIONWhat is the prevalence of defects in the Ca2+-signalling pathways mediating hyperactivation (calcium influx and store mobilization) among donors and sub-fertile patients and are they functionally significant, i.e. related to fertilization success at IVF?SUMMARY ANSWERThis study identifies, for the first time, the prevalence of Ca2+ store defects in sperm from research donors, IVF and ICSI patients. It highlights the biological role and importance of Ca2+ signalling (Ca2+ store mobilization) for fertilization at IVF.WHAT IS KNOWN ALREADYSperm motility and hyperactivation (HA) are important for fertility, mice with sperm incapable of HA are sterile. Recently, there has been significant progress in our knowledge of the factors controlling these events, in particular the generation and regulation of calcium signals. Both pH-regulated membrane Ca2+ channels (CatSper) and Ca2+ stores (potentially activating store-operated Ca2+ channels) have been implicated in controlling HA.STUDY DESIGN, SIZE, AND DURATIONThis was a prospective study examining a panel of 68 donors and 181 sub-fertile patients attending the Assisted Conception Unit, Ninewells Hospital Dundee for IVF and ICSI. Twenty-five of the donors gave a second sample (∼4 weeks later) to confirm consistency/reliability of the recorded responses. Ca2+ signalling was manipulated using three agonists, NH4Cl (activates CatSper via pH), progesterone (direct activation of CatSper channels, potentially enhancing mobilization of stored Ca2+ by CICR) and 4-aminopyridine (4-AP) (effect on pH equivalent to NH4Cl and mobilizes stored Ca2+). The broad-spectrum phosphodiesterase inhibitor 3-isobutyl-1-methyxanthine (IBMX), a potent activator of HA was also used for comparison. For patient samples, an aliquot surplus to requirements for IVF/ICSI treatment was examined, allowing direct comparison of Ca2+ signalling and motility data with functional competence of the sperm.MATERIALS, SETTING, METHODSThe donors and sub-fertile patients were screened for HA (using CASA) and changes in intracellular Ca2+ were assessed by loading with Fura-2 and measuring fluorescence using a plate reader (FluoStar).MAIN RESULTS AND THE ROLE OF CHANCEThe relative efficacy of the stimuli in inducing HA was 4-AP >> IBMX > progesterone. NH4Cl increased [Ca2+]i similarly to 4-AP and progesterone but did not induce a significant increase in HA. Failure of samples to generate HA (no significant increase in response to stimulation with 4-AP) was seen in just 2% of research donors but occurred in 10% of IVF patients (P = 0.025). All donor samples generated a significant [Ca2+]i increase when stimulated with 4-AP but 3.3% of IVF and 28.6% of ICSI patients failed to respond. Amplitudes of HA and [Ca2+]i responses to 4-AP were correlated with fertilization rate at IVF (P= 0.029; P = 0.031, respectively). Progesterone reliably induced [Ca2+]i responses (97% of donors, 100% of IVF patients) but was significantly less effective than 4-AP in inducing HA. Twenty seven per cent of ICSI patients failed to ...

Patch clamp studies of human sperm under physiological ionic conditions reveal three functionally and pharmacologically distinct cation channels

“…The fact that these substances can both activate ion channels raises the possibility that the channel underlying I Tail may contribute to the control of motility. Moreover, progesterone, which had no effect upon the sustained outward K + current but did augment I Tail , induces hyperactivation in a proportion of human sperm (Uhler et al , 1993; Fabbri et al , 1998; Teves et al , 2006; Sagare-Patil et al , 2012; Alasmari et al , 2013a, b). This response to progesterone may be critical for progress through the female tract and successful interaction with the egg and it is therefore interesting that spermatozoa from men with clinically identified fertility defects show that impaired activation by progesterone and 4-AP correlate well with reduced fertilization capacity (Alasmari et al , 2013a).…”

Mitochondria functionality and sperm quality

“…Calcium signalling and calcium store mobilisation have recently been shown to be important in Assisted Reproductive Technologies (ART) success, as responses are clearly different when patients are compared with sperm donors (Alasmari et al 2013). However, what role sperm mitochondria have in this process is open to question, although sperm mitochondria are known to uptake calcium, and have been hinted as a possible intracellular calcium store in human sperm (Costello et al 2009).…”

CatSper channel, sperm function and male fertility

“…An elevation in intracellular pH is the prerequisite for sperm hyperactivation. showed that extracellular application of cell permeant NH4Cl evokes elevation of intracellular pH, which stimulates Ca 2+ influx and induces sperm hyperactivation in bovine , but NH4Cl has negligible effect on human sperm hyperactivation (Alasmari et al, 2013).…”

“…In response to progesterone, CatSper mediates Ca 2+ influx, which is followed by Ca 2+ induced Ca 2+ release (CICR) that faclilitates the mobilization of Ca 2+ store at the sperm neck. The release of stored calcium is likely to be responsible for potent hyperactivation of human sperm perhaps by increased incidences of [Ca 2+ ]i oscillations (Alasmari et al, 2013). Moreover, what is the mechanism of Ca 2+ oscillations (repetitive transient [Ca 2+ ]i waves) during tail-tohead [Ca 2+ ]i propagation and what is the functional significance of unidirectional Ca 2+ propagation are further questions to be addressed?…”