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Submitted to: Sheep! Magazine
Publication Type: Popular Publication Publication Acceptance Date: 1/10/2007 Publication Date: 3/1/2007 Citation: Purdy, P.H. 2007. Preserving Ram Semen: New Science Shows it can be Stored for up to Two Days Before Freezing. Sheep! Magazine. p.40-41. Interpretive Summary: The effects of holding diluted ram semen at 5 degrees Celsius for up to 48 hours prior to cryopreservation were investigated. Semen from 6 rams was collected by electro-ejaculation in the autumn and again from 6 different rams in the spring. The sperm concentration and motility were determined. Samples were diluted at 23 degrees Celsius to 200 million sperm per mL in one-step, cooled to 5 degrees Celsius over 2 hours and maintained at 5 degrees Celsius for the duration of the experiments. Samples were loaded into French straws at 0, 24 or 48 hours after cooling, frozen in liquid nitrogen vapor, and plunged into liquid nitrogen for storage. After thawing, autumn and spring samples frozen after 0, 24, or 48 h of storage had similar percentages of motility, progressively motility, plasma membrane integrity and live acrosome reacted cells within a season. The quantity of sperm that bound to hen's egg membranes in autumn was not different across times but in spring the 0 h holding time had less sperm bound to hen's egg membranes compared to the 48 h holding time although the 24 h holding time was not different from the 0 or 48 h. This indicates that ram sperm can be held at 5 degrees Celsius for up to 48 hours prior to freezing with no injurious effects on motility, membrane integrity, or fertilizing potential as indicated by membrane binding ability. Technical Abstract: The effects of holding diluted ram semen at 5 degrees Celsius for up to 48 hours prior to cryopreservation were investigated. Semen from 6 rams was collected by electro-ejaculation in the autumn and again from 6 different rams in the spring. The sperm concentration and motility were determined using spectrophotometry and computerized automated semen analysis, respectively. Samples were diluted at 23 degrees Celsius to 200 x 106 cells per mL in one-step, cooled to 5 degrees Celsius over 2 hours and maintained at 5 degrees Celsius for the duration of the experiments. Samples were loaded into 0.5 mL French straws at 0, 24 or 48 hours after cooling, frozen in liquid nitrogen vapor, and plunged into liquid nitrogen for storage. After thawing, autumn samples frozen after 0, 24, or 48 h of storage exhibited similar percentages of motility (29, 31, 36%, respectively), progressively motility (16, 15, 17%, respectively), plasma membrane integrity (28, 35, 29%, respectively) and live acrosome reacted cells (0.4, 0.6, 0.8%, respectively; P > 0.05). In addition, the quantity of sperm that bound to hen's egg perivitelline membranes after being held at 5 degrees Celsius for 0, 24, or 48 h was not significantly different when the values were expressed as means of the quantity of sperm (155, 177, 106 sperm, respectively) or as the percentage of sperm inseminated (0.39, 0.49, 0.34, respectively; P > 0.05). Likewise, ram sperm collected in the spring and frozen at 0, 24 and 48 h after cooling had similar (P > 0.05) total motility (21, 25, 20%, respectively), progressive motility (14, 15, 11%, respectively), plasma membrane integrity (26, 33, 31%, respectively) and live acrosome reacted cells (3.7, 3.5, 3.2%, respectively; P > 0.05). The 0 h holding time had significantly less sperm bound to a hen's egg perivitelline membrane compared to the 48 h holding time (250 and 470 sperm, respectively) although the 24 h holding time was not different from the 0 or 48 h holding time (281 sperm; P < 0.05) but analysis of the proportion of the total sperm inseminated resulted in no significant differences observed (P > 0.05). These results indicate that ram sperm can be held at 5 degrees Celsius for up to 48 hours prior to freezing with no injurious effects on motility, membrane integrity, or fertilizing potential as indicated by membrane binding ability. |
