Recombinant human ZP3-induced sperm acrosome reaction: evidence for the involvement of T- and L-type voltage-gated calcium channels.
For successful fertilization mammalian spermatozoa must undergo the acrosome reaction (AR), an exocytotic event that allows this cell to penetrate the outer layer of the oocyte, the zona pellucida (ZP). Four glycoproteins (ZP1-ZP4) compose the human ZP, being ZP3 the physiological inductor of the AR. This process requires changes in intracellular Ca(2+) concentration ([Ca(2+)](i)) involving not fully understood mechanisms. Even in mouse sperm, the pharmacologically documented participation of voltage-gated Ca(2+) (Ca(V)) channels and store-operated channels (SOCs) in the ZP-induced AR is being debated. The situation in human sperm is even less clear due to the limited availability of human ZP. Here, we used recombinant human ZP3 (rhZP3) produced in baculovirus-infected Sf9 cells to investigate the involvement of Ca(V) channels in the human sperm AR. Our findings showed that Ni(2+) and mibefradil at concentrations that block T-type or Ca(V)3 channels, and nimodipine and diltiazem that block L-type or Ca(V)1 channels, significantly inhibited the rhZP3-initiated AR. On the other hand, the AR was insensitive to concentrations of omega-Agatoxin IVA, omega-Conotoxin GVIA and SNX-482 that block P/Q, N and R-type channels, respectively (Ca(V)2 channels). Our overall findings suggest that Ca(V)1 and Ca(V)3 channels participate in human sperm AR. Consistent with this, we detected in human sperm transcripts for the Ca(V)1 auxiliary subunits, alpha(2)delta, beta(1), beta(2) and beta(4), but not the neuronal specific isoforms beta(3) and gamma(2).
José
Omar
O
Departamento de Genética del Desarrollo y Fisiología Molecular, Instituto de Biotecnología, Universidad Nacional Autónoma de México (UNAM), Cuernavaca, Mexico.
Hernández-Hernández
Oscar
O
Chirinos
Mayel
M
González-González
María Elena
ME
Larrea
Fernando
F
Almanza
Angélica
A
Felix
Ricardo
R
Darszon
Alberto
A
Treviño
Claudia L
CL
D020101:Acrosome Reaction / Q000187:drug effects*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D020746:Calcium Channels, L-Type / Q000235:genetics / Q000502:physiology*; D020747:Calcium Channels, T-Type / Q000235:genetics / Q000502:physiology*; D002478:Cells, Cultured; D004527:Egg Proteins / Q000494:pharmacology*; D006801:Humans; D008297:Male; D008562:Membrane Glycoproteins / Q000494:pharmacology*; D020748:Mibefradil / Q000494:pharmacology; D009532:Nickel / Q000494:pharmacology; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins / Q000494:pharmacology*; D013094:Spermatozoa / Q000187:drug effects* / Q000502:physiology; D014158:Transcription, Genetic; D000072216:Zona Pellucida Glycoproteins; D020867:omega-Agatoxin IVA / Q000494:pharmacology; D020960:omega-Conotoxins / Q000494:pharmacology
D002121:Calcium Channel Blockers; D020746:Calcium Channels, L-Type; D020747:Calcium Channels, T-Type; D004527:Egg Proteins; D008562:Membrane Glycoproteins; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; C000606514:ZP1 protein, human; C000606517:ZP3 protein, human; D000072216:Zona Pellucida Glycoproteins; C000606519:Zp1 protein, mouse; C000606521:Zp3 protein, mouse; D020867:omega-Agatoxin IVA; D020960:omega-Conotoxins; D020748:Mibefradil; D009532:Nickel