Abstract
Whilst fertilizing capacity depends upon a K+ conductance (GK) that allows the spermatozoon membrane potential (Vm) to be held at a negative value, the characteristics of this conductance in human sperm are virtually unknown. We therefore studied the biophysical/pharmacological properties of the K+ conductance in spermatozoa from normal donors held under voltage/current clamp in the whole cell recording configuration. Our standard recording conditions were designed to maintain quasi-physiological, Na+, K+ and Cl− gradients. Experiments that explored the effects of ionic substitution/ion channel blockers upon membrane current/potential showed that resting Vm was dependent upon a hyperpolarizing K+ current that flowed via channels that displayed only weak voltage dependence and limited (∼7-fold) K+ versus Na+ selectivity. This conductance was blocked by quinidine (0.3 mM), bupivacaine (3 mM) and clofilium (50 µM), NNC55-0396 (2 µM) and mibefradil (30 µM), but not by 4-aminopyridine (2 mM, 4-AP). Progesterone had no effect upon the hyperpolarizing K+ current. Repolarization after a test depolarization consistently evoked a transient inward ‘tail current’ (ITail) that flowed via a second population of ion channels with poor (∼3-fold) K+ versus Na+ selectivity. The activity of these channels was increased by quinidine, 4-AP and progesterone. Vm in human sperm is therefore dependent upon a hyperpolarizing K+ current that flows via channels that most closely resemble those encoded by Slo3. Although 0.5 µM progesterone had no effect upon these channels, this hormone did activate the pharmacologically distinct channels that mediate ITail. In conclusion, this study reveals three functionally and pharmacologically distinct cation channels: Ik, ITail, ICatSper.
Highlights
Plasma membrane ion channels are central to the control of sperm function (Darszon et al, 1999; Lishko et al, 2011b) and, in particular, Ca2+entry via sperm cation channels (CatSper) is critical for several physiologically important processes, including hyperactivation, chemotaxis and the acrosome reaction (Lishko et al, 2011a, b; Strunker et al, 2011; Brenker et al, 2012)
Mouse and human spermatozoa normally display negative resting membrane potentials (Vm) that are dependent upon the activity of K+ channels, and the magnitude of this potential exerts a strong influence over Ca2+ influx since it determines the gating of CatSper and sets the driving force for Ca2+ entry through these channels
Recent experiments that directly compared the biophysical properties of mouse and human Slo3/ LRRC52 showed that the human channel complex could still pass hyperpolarizing K+ current when pHi was,7.0 (Leonetti et al, 2012), a result which accords well with the K+ currents which we describe in human spermatozoa themselves
Summary
Plasma membrane ion channels are central to the control of sperm function (Darszon et al, 1999; Lishko et al, 2011b) and, in particular, Ca2+. Entry via sperm cation channels (CatSper) is critical for several physiologically important processes, including hyperactivation, chemotaxis and the acrosome reaction (Lishko et al, 2011a, b; Strunker et al, 2011; Brenker et al, 2012). Mouse and human spermatozoa normally display negative resting membrane potentials (Vm) that are dependent upon the activity of K+ channels, and the magnitude of this potential exerts a strong influence over Ca2+ influx since it determines the gating of CatSper and sets the driving force for Ca2+ entry through these channels. Electrophysiological studies of mouse sperm led to the identification of the sperm K+ channel (KSper), a K+-permeable conductance whose activity was
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