Acrosomal integrity and capacitation are not influenced by sperm cryopreservation in the giant panda

Abstract

Sperm cryopreservation and artificial insemination are important management tools for giant panda breeding and the preservation of extant genetic diversity. This study examined the influence of freeze-thawing on sperm function, specifically capacitation. Sperm from nine giant pandas were assessed before and after rapid ( 2 40 and 2 100 8C/min) cryopreservation by incubation in HEPES-buffered Ham's F10 medium with and without the capacitation accelerators, 3-isobutyl-1-methylxanthine (IBMX) and dibutyryl cyclic AMP (dbcAMP). At 0, 3 and 6 h of exposure, aliquots were assessed for sperm motility traits and capacitation, defined as the proportion of sperm with intact acrosomes following exposure to solubilised zonae pellucidae (ursid or felid) or calcium ionophore subtracted from the proportion of sperm with intact acrosomes before exposure. Although mean6 S.E.M. sperm motility post-thaw (56.1 6 3.9% at 0 h) was less (P < 0.05) than pre-freeze (71.7 6 6.0%), there was no difference (P > 0.05) in the proportion of acrosome-intact sperm (fresh, 93.0 6 1.7% versus cryopreservedthawed, 81.7 6 4.7% at 0 h). Incidence of capacitation was greater (P < 0.05) in fresh sperm incubated with capacitation accelerators IBMX and dbcAMP (9 h: 50.9 6 1.1) compared with fresh sperm incubated without accelerators (9 h: 41.2 6 1.1%). Frozen-thawed sperm preincubated without accelerators underwent capacitation (49.6 6 1.1%) to a greater extent (P < 0.05) compared with these fresh counterparts. Thawed samples with (9 h: 45.9 6 1.4%) and without accelerators (9 h: 41.2 6 1.1%) did not differ (P > 0.05) during the 9-h incubation. We conclude that giant panda spermatozoa (1) undergo capacitation in vitro with or without chemical accelerators and (2) withstand a rapid cryopreservation protocol, including retaining normal acrosomal integrity and functional capacitation ability.

Flow cytometric cell cycle analysis of cultured brown bear fibroblast cells

“…Assisted reproductive technologies have been used to help in the preservation of endangered or threatened animals such as the African wild cat (Gomez et al, 2003) and giant panda (Han et al, 2003;Spindler et al, 2004;Hori et al, 2006). Although nuclear transfer raises controversial questions in its applications to wildlife conservation (Holt et al, 2004), the potential of this technology as a valuable tool for aiding in the conservation of some endangered and threatened animals should not be ignored.…”

Characteristics of Frozen-Thawed Spermatozoa Cryopreserved with Different Concentrations of Glycerol in Captive Japanese Black Bears (Ursus thibetanus japonicus)

“…Semen cryopreservation is one of the most important techniques for species conservation as a genetic resource. There are reports of semen cryopreservation in ursids, including giant pandas (Ailuropoda melanoleuca) [13,14,18], Hokkaido brown bears (Ursus arctos yesoensis) [7], and Japanese black bears [11,12]. However, although efforts to establish an optimal technique are important, no comparative studies of different glycerol concentrations of diluents for bear semen cryopreservation have been reported.…”

“…Characteristics of fresh semen collected by electroejaculation from Japanese black bears sperm [1]. Concentrations of 4.7% and 4-5% have been used for freezing the semen of closely related species, such as the brown bear [7] and giant panda [13,18], respectively. Our results corresponded to the conventional glycerol concentrations used in other ursids.…”

Cryopreservation of Brown Bear Skin Biopsies

“…Assisted reproductive technologies support the conservation efforts, 1 for example, to preserve the African wild cat 2 and giant panda, [3][4][5] In Spain, the Cantabric brown bear (Ursus arctos pyrenaicus) is an endangered population estimated at 100 individuals. Efforts are made to protect its declining environment and to preserve semen, cells, and somatic tissues.…”