Effects of arachnotoxin on intracellular pH and calcium in human spermatozoa.
OBJECTIVE
To determine the effect of arachnotoxin (ATx), a venom extracted from the Chilean spider Latrodectus mactans, on intracellular calcium ([Ca(2+)](i)) and pH (pH(i)) in capacitated human spermatozoa.


METHODS
Spermatozoa were collected from fertile adult men (n = 8). Mobile spermatozoa were collected by the "swim up" technique and stimulated with the crude extract of ATx and with progesterone (P).


METHODS
Hospital of the Federal University of São Paulo, São Paulo, Brazil.


METHODS
[Ca(2+)](i) was measured in fura2-AM-loaded spermatozoa, and pH(i) was measured in spermatozoa loaded with the pH-sensitive dye [(2',7')-bis (carboxymethyl)-(5,6)-carboxyfluorescein]-AM (BCECF).


RESULTS
The ATx and P induced a biphasic change in [Ca(2+)](i) consisting of a peak followed by a small but sustained elevation. The response to ATx was greatly reduced by pretreatment with P. The ATx caused intracellular acidification, whereas P induced alkalinization. Blockade of the NA(+)/H(+) exchanger with ethylisopropylamiloride (EIPA) sharply increased ATx-induced acidification.


CONCLUSIONS
Arachnotoxin increased [Ca(2+)](i) through the opening of calcium channels and release of calcium from intracellular stores. The ATx reduced pH(i) in human sperm, possibly by inhibiting the Na(+)/H(+) exchanger.
Romero
Fernando
F
Center of Reproductive Biotechnology, Department of Preclinical Sciences, Faculty of Medicine, Universidad de La Frontera, Temuco, Chile.
Cunha
Maria Adelaide
MA
Sanchez
Raul
R
Ferreira
Alice Teixeira
AT
Schor
Nestor
N
Oshiro
Maria Etsuko Miyamoto
ME
D020101:Acrosome Reaction / Q000187:drug effects*; D000328:Adult; D002118:Calcium / Q000378:metabolism*; D005452:Fluoresceins; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008297:Male; D013075:Sperm Capacitation / Q000187:drug effects*; D013094:Spermatozoa / Q000187:drug effects / Q000502:physiology*; D013111:Spider Venoms / Q000633:toxicity*
D005452:Fluoresceins; D013111:Spider Venoms; C043829:2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein; D002118:Calcium