Effects of osmotic pressure on motility, plasma membrane integrity and viability in fresh and frozen-thawed buffalo spermatozoa

Abstract

In the first experiment, osmotic pressure of semen and seminal plasma in a semen sample from each of the 20 mature NiliRavi buffalo bulls was determined. In the second experiment, effects of osmotic pressure on motility (%), plasma membrane integrity (%) and viability (%) in fresh and frozen-thawed semen samples from each of the seven mature Nili-Ravi buffalo bulls was determined. In the first experiment, seminal plasma was harvested by centrifuging semen at 400 3 g for 10 min at 378C and osmotic pressure was determined using an osmometer. In the second experiment, motility (%) was assessed in fresh and frozen-thawed (378C for 30 s) semen samples using a phase-contrast microscope (3400). Plasma membrane integrity (%) was determined by mixing 50 ml each of fresh and frozen-thawed semen with 500 ml of solution having an osmotic pressure of 50, 100, 150, 190 or 250 mOsm/l (hypotonic treatments of fructose 1 sodium citrate) and incubating at 378C for 1 h. Viability (%) of fresh and frozen-thawed spermatozoa before and after challenging them to osmotic pressure (hypotonic treatments) was assessed using supravital stain under a phase-contrast microscope (3400). In the first experiment, the mean 6 s.e. osmotic pressures of the buffalo semen and seminal plasma were 268.8 6 1.17 and 256.0 6 1.53 mOsm/l, respectively. In the second experiment, motility (%) decreased (P , 0.05) in frozen-thawed semen samples as compared with fresh semen (60.1 6 1.34 v. 81 6 1.57, respectively). The plasma membrane integrity (%) and magnitude of osmotic stress in fresh and frozen-thawed semen samples was higher (P , 0.05) at 50, 100, 150 and 190 mOsm/l as compared with 250 mOsm/l. Loss of viability (%) in fresh and frozen-thawed semen samples was higher (P , 0.05) at 50 mOsm/l (59% in fresh, 70% frozen thawed) as compared with other osmotic pressures, while it was lowest at 250 mOsm/l (4.1% for fresh, 9.7% frozen thawed). In conclusion, osmotic pressure of Nili-Ravi buffalo semen and seminal plasma is determined. Furthermore, variation in osmotic pressure below 250 mOsm/l is not favorable to fresh and frozen-thawed buffalo spermatozoa.

Optimal inclusion level of butylated hydroxytoluene in semen extender improves the quality of post-thawed canine sperm

“…A phase-contrast microscope (40×) was used for the assessment of percentage motility after placing a drop of thawed semen on a pre-warmed slide. Three observations were taken in order to attain a single point (Khan and Ijaz, 2008).…”

“…The hypo osmotic swelling test (HOST) was applied to assess the plasma membrane integrity of the spermatozoa as described earlier (Khan and Ijaz, 2008). 500 µl of hypo-osmotic solution (190 mOsm/l) was mixed with 50 µl of semen sample and the mixture was incubated at 37°C for 45 min.…”

A suplementação alimentar com ácidos graxos insaturados melhora a qualidade do sêmen criopreservado de búfalos (Bubalus bubalis) pós descongelação

“…Normalmente, as avaliações mais rotineiras realizadas para determinar a qualidade seminal englobam os testes de motilidade e morfologia espermática (MAZIERO et al, 2009), além de técnicas para avaliar a integridade e função da membrana plasmática do espermatozoide, como o teste de eosina-nigrosina e o teste hiposmótico (BARROS et al, 2007;SELVARAJU;GHOSH;RAVINDRA, 2009). De forma complementar, técnicas mais elaboradas foram desenvolvidas mais recentemente para aumentar o grau de precisão das avaliações andrológicas, como a análise computadorizada da motilidade espermática (CASA -Computer Assisted Sperm Analisys) (CONTRI et al, 2010;CONTRI et al, 2013;HUSSAIN;LESSARD;ANZAR, 2013) e as técnicas baseadas no uso de sondas fluorescentes (CELEGHINI et al, 2007;KOONJAENAK et al, 2007).…”

“…Esta ferramenta não somente pode ser utilizada para diminuir ou retirar a subjetividade do método convencional de avaliação da motilidade, mas também para investigar a cinética detalhada dos espermatozoides (MANDAL; BAGPAUL; GUPTA, 2003). Poucos estudos foram realizados com sêmen bubalino com o propósito de avaliar o comportamento da cinética espermática após os processos de congelação e descongelação (RASUL; AHMAD; ANZAR, 2000; RASUL; AHMAD; ANZAR, 2007;KOONJAENAK et al, 2007;ANIL KUMAR et al, 2011).…”

Evaluation of postthaw sperm parameters and fertility of Cholistani service bulls

“…Spermatozoa motility was assessed under phase contrast microscope (10), whereas viability was assessed through the vital staining technique (11). For the assessment of spermatozoa morphology and acrosomal integrity, each thawed semen sample was mixed with 1% formaldehyde citrate.…”