title: Voltage-dependent calcium influx in human sperm assessed by simultaneous optical detection of intracellular calcium and membrane potential.
abstract: There are several physiological and pharmacological evidences indicating that opening of voltage dependent calcium channels play a crucial role in the induction of the acrosome reaction in mammalian sperm. In mature sperm, physiological inductors of the acrosome reaction such as ZP3, a zona pellucida protein, and the steroid hormone progesterone, induce depolarization and calcium influx, which are required for the acrosome reaction. In this paper, we describe a voltage-dependent calcium influx present in human sperm. We report an experimental procedure that allows measurement of intracellular calcium and membrane potential simultaneously using the fluorescent dyes DiSC3(5) and Fura-2. We found that in human uncapacitated sperm, depolarization induces a nifedipine-insensitive calcium influx that, in most cases, was transient. Calcium influx was observed in the range of -60 to -15 mV (the range tested). At resting membrane potential (around -40 mV), potassium addition depolarized and induced calcium influx, but when the depolarization was preceded by a hyperpolarization (induced with valinomycin), calcium influx was remarkably enhanced, suggesting that at -40 mV, channels are in a putative inactivated state. When sperm was incubated in medium without calcium, calcium restoration caused calcium influx that depended on voltage, and decayed between 1 and 2 min after depolarization. Unlike ram, mouse or bovine sperm, in which an alkalinization is required to induce calcium influx with potassium, the voltage-dependent calcium influx observed in human sperm did not require an increase in internal or external pH. However, we observed that ammonium, which increases intracellular pH, enhanced the voltage-dependent calcium influx about 90%. Furthermore, depolarization by itself caused a small increase in intracellular pH suggesting that pH can be regulated by membrane potential in human sperm.
authors.affiliation: Departamento de Farmacología, Facultad de Medicina, Universidad Nacional Autónoma de México, D.F.
authors.forename: L
authors.initials: L
authors.lastname: Linares-Hernández
authors.forename: A M
authors.initials: AM
authors.lastname: Guzmán-Grenfell
authors.forename: J J
authors.initials: JJ
authors.lastname: Hicks-Gomez
authors.forename: M T
authors.initials: MT
authors.lastname: González-Martínez
mesh_terms: D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000187:drug effects / Q000502:physiology*; D002138:Calibration; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007424:Intracellular Fluid / Q000378:metabolism*; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects; D009532:Nickel / Q000494:pharmacology; D009543:Nifedipine / Q000494:pharmacology; D011189:Potassium Chloride / Q000494:pharmacology; D013075:Sperm Capacitation / Q000187:drug effects; D013094:Spermatozoa / Q000378:metabolism / Q000502:physiology*; D014634:Valinomycin / Q000494:pharmacology
chemical_list: D015220:Calcium Channels; D014634:Valinomycin; D011189:Potassium Chloride; D009532:Nickel; D009543:Nifedipine; D002118:Calcium