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J. L. Courtens, L. Plöen, and M. Loir

Summary. Protamine was specifically demonstrated in spermatids and spermatozoa of the boar by immunoelectron microscopy, using anti-boar or anti-ram protamine antisera, and three different direct or indirect labelling techniques. The two isomers of the protamine could not be labelled separately. The protamine is present in the cytoplasm of elongating spermatids and it enters the nuclei throughout the elongation process after possible storage in the cytoplasm or in the nuclear envelope of spermatids, or both. These findings differ from previous observations in other species.

Keywords: spermiogenesis; spermatozoa; protamine; immunocytochemistry; electron microscopy; boar

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J. L. Courtens, H. Ekwall, M. Paquignon, and L. Plöen

Summary. Boar semen was analysed by electron microscopy coupled to image analysis and X-ray energy dispersive spectroscopy, during the usual process for freezing and thawing in field conditions. Freeze–substitution and freeze–quenching permitted recording of real or potential intracellular ice before, during, and after freezing. Heads and flagella displayed two different osmotic properties before freezing. Heads were dehydrated progressively before and during freezing, while flagella were hydrated before freezing and were only dehydrated during freezing. All parts of the thawed cells were rehydrated. Ice crystal damage was mostly present in frozen mitochondria and axonemes and the acrosomes were strongly affected by thawing. The total amounts of Na, Cl, Ca, K, Mg, and Zn per cell were only elevated in frozen and thawed midpieces while the heads were permeable both to water and elements at that time.

Keywords: boar; spermatozoa; freezing; water; element concentrations; electron microscopy; image analysis; X-ray spectrophotometry

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H. Rodriguez-Martinez, J. L. Courtens, U. Kvist, and L. Plöen

Summary. Protamine was specifically demonstrated in boar spermatozoa collected from the rete testis, caput, corpus and cauda epididymidis and the ejaculate by immunoelectron microscopy, using anti-boar or anti-ram protamine antisera and an indirect post-embedding immunogold technique. Spermatozoa from all collection sites stained after incubation although with different degrees of labelling. Controls were negative. Labelling increased from the rete testis towards the epididymal corpus, where it was most intense, decreasing sharply thereafter. The weakest binding of the assayed antibodies was obtained in the ejaculated spermatozoa but it could be reversed by in-vitro induction of chromatin decondensation with sodium dodecyl sulphate and the metal-chelating EDTA. The finding of a significant decrease in the immunolabelling detected from the corpus epididymidis onwards indicates a critical point for the interaction between DNA and the protamines in boar spermatozoa during the epididymal maturation.

Keywords: spermatozoa; epididymis; protamine; immunocytochemistry; electron microscopy; boar