Disappearance of the PHA-E Lectin Binding Site on the Surface of Ejaculated Sperm and Sperm Capacitation in the Dog

Abstract

ABSTRACT. Ejaculated semen and cross sections of the cauda epididymides collected from 3 normal dogs were smeared or stamped on glass slides, and the sperm on the slides were stained with 7 different FITC-lectins (Con A, DBA, GS-1, PHA-E, PSA, UEA-1, WGA) to examine the relation between sperm-binding glycoprotein derived from the canine prostate and sperm capacitation. The only lectin that stained the ejaculated sperm but not the cauda epididymal sperm was PHA-E. The sperm ejaculated by 5 other dogs were incub ated for 4 hr in fluid flushed from the uterine horns or oviducts of estrous bitches, and then the percentages of actively motile sperm and hyperactivated sperm (HA-sperm) were determined. The percentages of PHA-E-labeled sperm and sperm labeled with fluoresceinated Ca indicator to assess the influx of Ca into the sperm were also evaluated. The mean percentages of actively motile sperm, HA-sperm, and Ca-labeled sperm after 4 hr of incubation in the uterine flush fluid and oviductal flush fluid were significantly higher tha n in control medium (P<0.05, 0.01), but the mean percentages of PHA-E-labeled sperm were lower (both P<0.01). The percentages of PHA-E-unlabeled sperm correlated with the percentages of both HA-sperm and Ca-labeled sperm (r 2 =0.787 and 0.812, respectively). The results indicate that loss of the glycoprotein secreted by the canine prostate on the sperm surface induces the influx of Ca into the sperm, and then hyperactivation of the sperm. KEY WORDS: canine, capacitation, lectin, prostate, sperm.J. Vet. Med. Sci. 66 (5): [495][496][497][498][499][500] 2004 It has been reported that the surface of the plasma membrane of mammalian sperm becomes coated with various glycoproteins secreted by the epididymal epithelium during sperm transit through the epididymis [9,11], and some of the glycoproteins on the sperm surface are thought to induce sperm maturation in the epididymal duct [10,17,29]. It is known that the glycoproteins produced by the prostates and seminal vesicles are contained in the semen [2,5,24] and that some of them act as decapacitation factors [2,24]. The decapacitation factors on the surface of the sperm are lost as a result of the action of proteolytic enzymes in the uterus and oviduct [5,6,19], and sperm capacitation is then promoted by some components of oviductal fluid, e.g., glycosaminoglycans [11,15,22]. Sperm capacitation is induced by an influx of Ca into the cytoplasm of the sperm [23,25,27]. Techniques utilizing many different fluorescein-isothiocyanate (FITC)-labeled lectins have revealed the characteristics of the glycoproteins coating the surface of the sperm in many species [9,[16][17][18]29], but there have been few reports on the relation between sperm-binding glycoproteins and sperm capacitation in the dog [3,7]. In the present study, canine epididymal sperm and ejaculated sperm were stained with 7 different FITC-lectins to examine for the presence of sperm-binding glycoproteins derived from canine prostate in the seminal fluid and to assess the relation bet...

Effects of Hyaluronic Acid on Ca²⁺ Influx, Lactate Dehydrogenase Activity, and Cyclic AMP Synthesis in Canine Ejaculated Sperm during <i>In Vitro</i> Capacitation

“…Evaluation of Ca 2+ influx into sperm cytoplasm: Ca 2+ influx into the cytoplasm was assessed 4 hr after the start of incubation by staining with a fluoresceinated Ca 2+ indicator (Calcium Crimson, Molecular Probes, Inc., Oregon, U.S.A.) according to the method described previously [22]. The percentage of sperm with fluoresceinated heads (Ca 2+ -labeled sperm) was estimated by examining 300 sperm with a flurescence microscope (BX60, Olympus Corporation, Tokyo).…”