Induction of capacitation and the acrosome reaction of boar spermatozoa by L-arginine and nitric oxide synthesis associated with the anion transport system

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Abstract

The aim of this study was to determine the effect of L-arginine on nitric oxide (NO) synthesis, capacitation and acrosome reaction of boar spermatozoa. Ejaculated boar spermatozoa were washed and then cultured in a bicarbonate: CO2-buffered medium, modified NCSU-37, for 2 h. At the end of the culture, the status of spermatozoa was determined. The presence of (0.1-2.0 mmol l-1) L-arginine in the culture medium induced an acrosome reaction determined fluorescein isothiocyanate-peanut agglutinin (FITC-PNA) and increased intracellular NO content, as quantified by a fluorescent indicator, diaminofluorescein-2 diacetate (DAF-2 DA). This stimulatory effect of L-arginine was neutralized by supplementation with an NO synthase inhibitor, Nω-nitro-L-arginine methyl ester (1 mmol l-1). However, the inactive enantiomorph, Nω-nitro-D-arginine methyl ester, did not affect the stimulatory effect of L-arginine. These results indicate that L-arginine induces an acrosome reaction through the NO signal pathway in boar spermatozoa. Furthermore, the stimulatory effect of L-arginine was inhibited in the presence of an anion transport inhibitor, 4-acetamido-4′-isothiocyanostilbene-2.2′-disulphonic acid (SITS; 0.1 mmol l-1), whereas any responses of spermatozoa to caffeine were not inhibited by SITS. A stimulatory effect of L-arginine on capacitation and acrosome reaction of spermatozoa was also observed in modified NCSU37 medium by using a chlortetracycline fluorescence assay, but not in supplemented bicarbonate-free Tris-buffered medium. These results indicate that the presence of L-arginine induces nitric oxide synthesis and stimulates capacitation and acrosome reaction of boar spermatozoa only when active sperm anion transport is present as a result of bicarbonate supplementation.

Original languageEnglish
Pages (from-to)857-864
Number of pages8
JournalReproduction
Volume124
Issue number6
Publication statusPublished - Dec 1 2002

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Acrosome Reaction
Anions
Arginine
Spermatozoa
Nitric Oxide
Bicarbonates
4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid
Sperm Transport
Chlortetracycline
Caffeine
Nitric Oxide Synthase
Culture Media
Signal Transduction
Fluorescence
Acids

ASJC Scopus subject areas

  • Obstetrics and Gynaecology
  • Cell Biology
  • Endocrinology
  • Embryology

Cite this

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title = "Induction of capacitation and the acrosome reaction of boar spermatozoa by L-arginine and nitric oxide synthesis associated with the anion transport system",
abstract = "The aim of this study was to determine the effect of L-arginine on nitric oxide (NO) synthesis, capacitation and acrosome reaction of boar spermatozoa. Ejaculated boar spermatozoa were washed and then cultured in a bicarbonate: CO2-buffered medium, modified NCSU-37, for 2 h. At the end of the culture, the status of spermatozoa was determined. The presence of (0.1-2.0 mmol l-1) L-arginine in the culture medium induced an acrosome reaction determined fluorescein isothiocyanate-peanut agglutinin (FITC-PNA) and increased intracellular NO content, as quantified by a fluorescent indicator, diaminofluorescein-2 diacetate (DAF-2 DA). This stimulatory effect of L-arginine was neutralized by supplementation with an NO synthase inhibitor, Nω-nitro-L-arginine methyl ester (1 mmol l-1). However, the inactive enantiomorph, Nω-nitro-D-arginine methyl ester, did not affect the stimulatory effect of L-arginine. These results indicate that L-arginine induces an acrosome reaction through the NO signal pathway in boar spermatozoa. Furthermore, the stimulatory effect of L-arginine was inhibited in the presence of an anion transport inhibitor, 4-acetamido-4′-isothiocyanostilbene-2.2′-disulphonic acid (SITS; 0.1 mmol l-1), whereas any responses of spermatozoa to caffeine were not inhibited by SITS. A stimulatory effect of L-arginine on capacitation and acrosome reaction of spermatozoa was also observed in modified NCSU37 medium by using a chlortetracycline fluorescence assay, but not in supplemented bicarbonate-free Tris-buffered medium. These results indicate that the presence of L-arginine induces nitric oxide synthesis and stimulates capacitation and acrosome reaction of boar spermatozoa only when active sperm anion transport is present as a result of bicarbonate supplementation.",
author = "Hiroaki Funahashi",
year = "2002",
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language = "English",
volume = "124",
pages = "857--864",
journal = "Reproduction",
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N2 - The aim of this study was to determine the effect of L-arginine on nitric oxide (NO) synthesis, capacitation and acrosome reaction of boar spermatozoa. Ejaculated boar spermatozoa were washed and then cultured in a bicarbonate: CO2-buffered medium, modified NCSU-37, for 2 h. At the end of the culture, the status of spermatozoa was determined. The presence of (0.1-2.0 mmol l-1) L-arginine in the culture medium induced an acrosome reaction determined fluorescein isothiocyanate-peanut agglutinin (FITC-PNA) and increased intracellular NO content, as quantified by a fluorescent indicator, diaminofluorescein-2 diacetate (DAF-2 DA). This stimulatory effect of L-arginine was neutralized by supplementation with an NO synthase inhibitor, Nω-nitro-L-arginine methyl ester (1 mmol l-1). However, the inactive enantiomorph, Nω-nitro-D-arginine methyl ester, did not affect the stimulatory effect of L-arginine. These results indicate that L-arginine induces an acrosome reaction through the NO signal pathway in boar spermatozoa. Furthermore, the stimulatory effect of L-arginine was inhibited in the presence of an anion transport inhibitor, 4-acetamido-4′-isothiocyanostilbene-2.2′-disulphonic acid (SITS; 0.1 mmol l-1), whereas any responses of spermatozoa to caffeine were not inhibited by SITS. A stimulatory effect of L-arginine on capacitation and acrosome reaction of spermatozoa was also observed in modified NCSU37 medium by using a chlortetracycline fluorescence assay, but not in supplemented bicarbonate-free Tris-buffered medium. These results indicate that the presence of L-arginine induces nitric oxide synthesis and stimulates capacitation and acrosome reaction of boar spermatozoa only when active sperm anion transport is present as a result of bicarbonate supplementation.

AB - The aim of this study was to determine the effect of L-arginine on nitric oxide (NO) synthesis, capacitation and acrosome reaction of boar spermatozoa. Ejaculated boar spermatozoa were washed and then cultured in a bicarbonate: CO2-buffered medium, modified NCSU-37, for 2 h. At the end of the culture, the status of spermatozoa was determined. The presence of (0.1-2.0 mmol l-1) L-arginine in the culture medium induced an acrosome reaction determined fluorescein isothiocyanate-peanut agglutinin (FITC-PNA) and increased intracellular NO content, as quantified by a fluorescent indicator, diaminofluorescein-2 diacetate (DAF-2 DA). This stimulatory effect of L-arginine was neutralized by supplementation with an NO synthase inhibitor, Nω-nitro-L-arginine methyl ester (1 mmol l-1). However, the inactive enantiomorph, Nω-nitro-D-arginine methyl ester, did not affect the stimulatory effect of L-arginine. These results indicate that L-arginine induces an acrosome reaction through the NO signal pathway in boar spermatozoa. Furthermore, the stimulatory effect of L-arginine was inhibited in the presence of an anion transport inhibitor, 4-acetamido-4′-isothiocyanostilbene-2.2′-disulphonic acid (SITS; 0.1 mmol l-1), whereas any responses of spermatozoa to caffeine were not inhibited by SITS. A stimulatory effect of L-arginine on capacitation and acrosome reaction of spermatozoa was also observed in modified NCSU37 medium by using a chlortetracycline fluorescence assay, but not in supplemented bicarbonate-free Tris-buffered medium. These results indicate that the presence of L-arginine induces nitric oxide synthesis and stimulates capacitation and acrosome reaction of boar spermatozoa only when active sperm anion transport is present as a result of bicarbonate supplementation.

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