PARENT SESSION
SLIDE SESSION 3: REGULATION OF SPERM FUNCTION
Chairs: Gary Killian, Marc-Andre Sirard, TanYa Gwathmey (Trainee)
Arts Hall 026
2:30 PM-4:30 PM
20
THE GLYCINE RECEPTOR IS INVOLVED THE ZP3-INITIATED ACROSOME REACTION OF HUMAN SPERM.
Son, Jung-Ho1, Bray, Chris1, Harris, Jeffrey2, Meizel, Stanley1, 1 2
ABSTRACT- We have previously demonstrated a role for the sperm glycine receptor (GlyR) in the pig and mouse sperm acrosome reaction (AR) initiated by the egg zona pellucida (ZP). The ZP protein ZP3 (ZPC) is known to initiate the AR. The aim of the present study was: 1) to determine whether a particular recombinant ZP3 utilized a G protein-mediated mechanism similar to that of native ZP for AR initiation; 2) to determine whether the human sperm GlyR is also involved in that event. Purified recombinant human ZP3 (rhuZP3) was prepared as described by Harris et al., 1999 and tested for biological activity on 24 hr-capacitated sperm using the ConA lectin-FITC AR assay. Results are shown as mean % AR ± SEM. Incubation with rhuZP3 (20 min; 100 ng/ml) was shown to stimulate AR (rhuZP3 32.1 ± 4.90 % vs PBS control 18.1 ± 3.5; n=6; p <0.05). Pertussis toxin (a classical Gi protein inhibitor) is a known blocker of the ZP-initiated AR. Pre-incubation with pertussis toxin (100 ng/ml; 3 hr), completely inhibited the effect of rhuZP3 (16.4 ± 1.9 %; n=6; p<0.05), suggesting the rhuZP3 was acting through the same mechanism as solubilized native ZP. Sperm were also preincubated with either an antibody against the alpha1 subunit of the human neuronal GlyR (GlyR Ab; 3 mg/ml; 30 min) or the classical GlyR inhibitor, strychnine (ST; 50 nM; 10 min) prior to AR initiation with rhuZP3. Both GlyR Ab and ST inhibited the rhuZP3 AR (GlyR Ab + rhuZP3 22.0 ± 2.3 % vs PBS + rhuZP3 30.0 ± 2.9 %; n=3; p<0.05; ST + rhuZP3 20.5 ± 2.3 vs PBS + rhuZP3 30.3 ± 2.8; n=3; p<0.05). These results indicated the GlyR plays an important role in the human sperm AR initiated by ZP and further support the view that AR initiation by rhuZP3 and native ZP utilize the same mechanisms. Funded by NIH grant HD-33368 to SM
KEY WORDS: recombinant ZP3, Glycine receptor, acrosome reaction, human sperm