In vitro capacitating effect of gamma-aminobutyric acid in ram spermatozoa.
We have evaluated the capacitating effect of gamma-aminobutyric acid (GABA) in ram spermatozoa in vitro, in a chemically defined medium, by means of the chlortetracycline (CTC) binding assay. Semen from adult Australian Merino rams was collected in an artificial vagina; spermatozoa were washed once in modified Biggers, Whitten, and Wittingham medium (m-BWW), without BSA or serum, and incubated in m-BWW alone or in m-BWW containing GABA, GABA agonists, or antagonists for 2 h at 38.5 degrees C under 5% CO2 in air. Samples were taken for assessment of CTC binding pattern or were further incubated for 15 min in the presence of 5 microM calcium ionophore A23187. Acrosomal exocytosis was evaluated by Pisum sativum agglutinin binding. Addition of GABA to the incubation medium resulted in a concentration-dependent increase in the percentage of CTC forms II and III, corresponding to mid-capacitated and capacitated spermatozoa, respectively. The effect was marginally significant at 1 microM and maximal at 20 microM. The action of 20 microM GABA was mimicked by the GABAB-receptor agonist, muscimol, but not by the GABAA-receptor agonist, baclofen, and completely blocked by the GABAA-receptor antagonists, bicuculline and picrotoxin, which lacked effect per se. In a separate set of experiments, incubation of spermatozoa with GABA at a concentration of 1 microM, which was insufficient to stimulate sperm capacitation, together with the neuroactive steroid allopregnanolone (1 microM) provoked a capacitating effect similar to that achieved by 20 microM GABA alone. These results show that GABA has a capacitating action on ram spermatozoa through a GABAA receptor-mediated mechanism.
de las Heras
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Centro de Investigaciones Reproductivas Perez Companc, Buenos Aires, Argentina. cirpc@datamarkets.com.ar
Valcarcel
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A
Perez
L J
LJ
D000177:Acrosome / Q000187:drug effects / Q000502:physiology*; D000704:Analysis of Variance; D000818:Animals; D001418:Baclofen / Q000494:pharmacology; D000001:Calcimycin / Q000494:pharmacology; D002751:Chlortetracycline / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005089:Exocytosis / Q000187:drug effects; D018755:GABA Agonists / Q000494:pharmacology; D018756:GABA Antagonists / Q000494:pharmacology; D066298:In Vitro Techniques; D008297:Male; D009118:Muscimol / Q000494:pharmacology; D018696:Neuroprotective Agents / Q000494:pharmacology; D011280:Pregnanolone / Q000494:pharmacology; D017430:Prostate-Specific Antigen / Q000032:analysis; D011963:Receptors, GABA-A / Q000502:physiology; D018080:Receptors, GABA-B / Q000502:physiology; D012756:Sheep; D013075:Sperm Capacitation / Q000187:drug effects*; D013094:Spermatozoa / Q000187:drug effects / Q000502:physiology*; D005680:gamma-Aminobutyric Acid / Q000494:pharmacology*
D018755:GABA Agonists; D018756:GABA Antagonists; D018696:Neuroprotective Agents; D011963:Receptors, GABA-A; D018080:Receptors, GABA-B; D009118:Muscimol; D000001:Calcimycin; D005680:gamma-Aminobutyric Acid; D011280:Pregnanolone; D017430:Prostate-Specific Antigen; D001418:Baclofen; D002751:Chlortetracycline