Bicarbonate dependence of cAMP accumulation induced by phorbol esters in hamster spermatozoa.
Phorbol esters stimulate cyclic adenosine 3',5'-monophosphate (cAMP) accumulation in hamster spermatozoa under conditions for in vitro capacitation. The 20-50-fold elevation of cAMP levels induced by 1 microM phorbol 12-myristate 13-acetate (PMA) in spermatozoa depends on the presence of sodium bicarbonate in the medium (ED50: 15 mM) and it is independent of extracellular pH. Sodium bicarbonate stimulates adenylate cyclase activity in membrane preparations by 4-fold (ED50: 40 mM). After solubilization, the bicarbonate-sensitive moiety elutes as a single peak of 55 kDa in a gel filtration column. Blockers of bicarbonate chloride antiporters diisothiocyanate stilbene 2,2'-disulfonic acid (DIDS) or acetamido 4'-isothiocyanate stilbene 2,2'-disulfonic acid (SITS) inhibit the bicarbonate dependent PMA effect on cAMP in living spermatozoa (ED50: 100 microM). Maximal (85%) inhibition in cAMP accumulation is observed at 1 mM. Motility is inhibited only at high concentrations of the blockers. Pretreatment of living cells with 1 mM DIDS does not affect membrane adenylate cyclase activity which remains responsive to bicarbonate. These results suggest that controlled transport of bicarbonate through the sperm plasma membrane could be associated to the regulation of cAMP synthesis.
Visconti
P E
PE
Instituto de Biología y Medicina Experimental, Buenos Aires, Argentina.
Muschietti
J P
JP
Flawia
M M
MM
Tezon
J G
JG
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D000262:Adenylyl Cyclases / Q000378:metabolism*; D000818:Animals; D001639:Bicarbonates / Q000494:pharmacology*; D001692:Biological Transport; D006224:Cricetinae; D000242:Cyclic AMP / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D004822:Epididymis / Q000187:drug effects / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D008297:Male; D015240:Phorbol 12,13-Dibutyrate / Q000493:pharmacokinetics / Q000494:pharmacology*; D012964:Sodium / Q000494:pharmacology*; D017693:Sodium Bicarbonate; D013075:Sperm Capacitation / Q000187:drug effects*; D013094:Spermatozoa / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*
D001639:Bicarbonates; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; D015240:Phorbol 12,13-Dibutyrate; D017693:Sodium Bicarbonate; D012964:Sodium; D000242:Cyclic AMP; D000262:Adenylyl Cyclases; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid