Depolarization of sperm membrane potential is a common feature of men with subfertility and is associated with low fertilization rate at IVF: Potassium channel dysfunction in human spermatozoa

Study question: Are significant abnormalities in outward (K+) conductance and resting membrane potential (Vm) present in the spermatozoa of patients undertaking IVF and Intra Cytoplasmic Sperm Injection (ICSI) and if so, what is their functional effect on fertilization success? Summary answer: Negligible outward conductance (≈5% of patients) or an enhanced inward conductance (≈4% of patients), both of which caused depolarisation of Vm, were associated with a low rate of fertilisation following IVF. What is known already: Sperm-specific potassium channel knockout mice are infertile with defects in sperm function suggesting that these channels are essential for fertility. These observations suggest that malfunction of K+ channels in human spermatozoa might contribute significantly to the occurrence of sub-fertility in men. However, remarkably little is known of the nature of K+ channels in human spermatozoa or the incidence and functional consequences of K+ channel defects. Study design, size, and duration: Spermatozoa were obtained from healthy volunteer research donors and sub fertile IVF and ICSI patients attending a hospital assisted reproductive techniques clinic between May 2013 and December 2015. In total 40 IVF patients, 40 ICSI patients and 26 normozoospermic donors took part in the study. Participants/materials, setting, methods: Samples were examined using electrophysiology (whole cell patch clamping). Where abnormal electrophysiological characteristics were identified, spermatozoa were further examined for Ca2+ influx induced by progesterone and penetration into viscous media if sufficient sample was available. Full exome sequencing was performed to specifically evaluate KCNMA1, KCNU1 and LRRC52 genes and others associated with K+ signalling. In IVF patients comparison with fertilisation rates was done to assess the functional significance of the electrophysiological abnormalities. Main results and the role of chance: Patch clamp electrophysiology was used to assess outward (K+) conductance and resting membrane potential (Vm) and signalling/motility assays were used to assess functional characteristics of sperm from IVF and ICSI patient samples. Mean Vm and outward membrane conductance in sperm from IVF and ICSI patients were not significantly different to those of control (donor) sperm prepared under the same conditions, but variation between individuals was significantly greater (P<0.02) with a large number of outliers (>25%). In particular, in ≈10% of patients (7/81) we observed either a negligible outward conductance (4 patients) or an enhanced inward current (3 patients), both of which caused depolarisation of Vm. Analysis of clinical data from the IVF patients showed significant association of depolarised Vm (≥0 mV) with low fertilisation rate (P=0.012). Spermatozoa with electrophysiological abnormities (conductance and Vm) responded normally to progesterone with elevation of [Ca2+]i and penetration of viscous medium, indicating retention of cation channel of sperm (CatSper) channel function. Limitations, reasons for caution: For practical, technical, ethical and logistical reasons we could not obtain sufficient additional semen samples from men with conductance abnormalities to establish the cause of the conductance defects. Full exome sequencing was only available in 2 men with conductance defects Wider implications of the findings: These data add significantly to the understanding of the role of ion channels in human sperm function and its impact on male fertility. Impaired potassium channel conductance (Gm) and/or Vm regulation is both common and complex in human spermatozoa and importantly is associated with impaired fertilization capacity when the Vm of cells is completely depolarized.