title: Cryopreservation of Atlantic salmon Salmo salar sperm: effects on sperm physiology.
abstract: The objective of this study was to determine the effect of freezing on the function in Atlantic salmon Salmo salar spermatozoa. The semen was frozen in Cortland's medium + 1.3M dimethyl sulphoxide + 0.3M glucose + 2% bovine serum albumin (final concentration) in a ratio of 1:3 (semen:cryoprotectant) as the treatment (T) and fresh semen as the control (F). Straws of 0·5 ml of sperm suspension were frozen in 4 cm of N2 L. They were thawed in a thermoregulated bath (40° C). After thawing, the percentage of spermatozoa with fragmented DNA [transferase dUTP (deoxyuridine triphosphate) nick-end labelling (TUNEL)], plasma membrane integrity (SYBR-14/PI) and mitochondrial membrane potential (ΔΨMMit, JC-1) were evaluated by flow cytometry and motility was evaluated by optical microscope under stroboscopic light. The fertilization rates of the control and treatment semen were tested at a sperm density of 1·5 × 10(7) spermatozoa oocyte(-1) , by observation of the first cleavages after 16 h incubation at 10° C. In the cryopreserved semen (T), the mean ± s.d. DNA fragmentation was 4·8 ± 2·5%; plasma membrane integrity 75·2 ± 6·3%; mitochondrial membrane potential 51·7 ± 3·6%; motility 58·5 ± 5·3%; curved line velocity (VCL ) 61·2 ± 17·4 µm s(-1) ; average-path velocity (VAP ) 50·1 ± 17·3 µm s(-1) ; straight-line velocity (VSL ) 59·1 ± 18·4 µm s(-1) ; fertilization rate 81·6 ± 1·9%. There were significant differences in the plasma membrane integrity, mitochondrial membrane potential, motility, fertilization rate, VCL , VAP and VSL compared with the controls (P < 0·05). Also the mitochondrial membrane potential correlated with motility, fertilization rate, VCL and VSL (r = 0·75; r = 0·59; r = 0·77 and r = 0·79, respectively; P < 0·05); and the fertilization rate correlated with VCL and VSL (r = 0·59 and r = 0·55, respectively).
authors.affiliation: School of Aquaculture, Catholic University of Temuco, Temuco, Chile.|Departamento de Ingeniería Química, Facultad de Ingeniería y Ciencias, Universidad de La Frontera, Temuco, Chile.
authors.forename: E
authors.initials: E
authors.lastname: Figueroa
authors.affiliation: School of Aquaculture, Catholic University of Temuco, Temuco, Chile.
authors.forename: I
authors.initials: I
authors.lastname: Valdebenito
authors.affiliation: BIOREN-Center for Biotechnology in Reproduction, La Frontera University, Temuco, Chile.
authors.forename: O
authors.initials: O
authors.lastname: Merino
authors.affiliation: School of Aquaculture, Catholic University of Temuco, Temuco, Chile.
authors.forename: A
authors.initials: A
authors.lastname: Ubilla
authors.affiliation: BIOREN-Center for Biotechnology in Reproduction, La Frontera University, Temuco, Chile.
authors.forename: J
authors.initials: J
authors.lastname: Risopatrón
authors.affiliation: Departamento de Ingeniería Química, Facultad de Ingeniería y Ciencias, Universidad de La Frontera, Temuco, Chile. jorge.farias@ufrontera.cl.
authors.forename: J G
authors.initials: JG
authors.lastname: Farias
mesh_terms: D000818:Animals; D015925:Cryopreservation; D003451:Cryoprotective Agents; D005306:Fertilization; D008297:Male; D018613:Microscopy, Confocal; D009865:Oocytes; D009930:Organic Chemicals; D020317:Salmo salar; D012661:Semen; D012662:Semen Preservation; D013076:Sperm Count; D013081:Sperm Motility; D013094:Spermatozoa / Q000502:physiology
chemical_list: D003451:Cryoprotective Agents; D009930:Organic Chemicals; C093207:SYBR-14
keywords: fertility; fish; sperm function; spermatozoa freezing