Hypoosmotic Swelling Test in Alpaca (Vicugna pacos) Epididymal Spermatozoa

Abstract

The present study intended to develop the hypoosmotic swelling (HOS) test in alpaca for its use in epididymal spermatozoa, to evaluate the membrane functional integrity and determine an appropriate hypoosmotic solution and whether the incubation time of 15 or 60 min is sufficient for the execution of the test. Hypoosmotic solutions (HS) with the following concentrations were used: 50, 100, 150, 200 and 275 mOsm/kg of sodium citrate tribasic dihydrate and d-fructose. Ten microlitres of epididymal sperm sample was mixed in 150 μL of the respective HS and incubated for 15 or 60 min at 38°C. From the proportion of reacted (swollen) spermatozoa, the 150 mOsm/kg HS was the most sensitive (p < 0.05). The exposure times (15 and 60 min) did not have significant differences (p > 0.05) in the proportion of both strong- and total-coiled sperm tails. In conclusion, 150 mOsm/kg HS and 15 min exposure time are optimal to evaluate the plasma membrane functional integrity through the HOS test in alpaca epididymal spermatozoa.

Epididymis incision as a method to collect epididymal sperm cells in alpacas

“…The volumes (1-4 mL) and temperatures (35-37 • C) of the media used, as well as the incubation time vary (Abraham et al, 2016;Kershaw-Young & Maxwell, 2011;Mamani-Mango et al, 2019;Vet Med Sci. 2022;8:157-163. wileyonlinelibrary.com/journal/vms3 Morton et al, 2010b;Morton et al, 2007;Vasquéz et al, 2012). With this method of extraction, Morton et al (2010b;2007) achieved a success rate of 100% (n = 10 each) in three experiments and 90.90%…”

“…No information was provided on the success of the extraction. A study by Vasquéz et al (2012) used 28 epididymides, but the success rate was not stated. Mamani-Mango et al (2019) compared sperm extraction by crushing the epididymis with retrograde flushing.…”

“…In alpaca, the prepared and cleaned epididymis is usually divided into small pieces and placed in liquid to allow the sperm to float out. The volumes (1–4 mL) and temperatures (35–37°C) of the media used, as well as the incubation time vary (Abraham et al., 2016 ; Kershaw‐Young & Maxwell, 2011 ; Mamani‐Mango et al., 2019 ; Morton et al., 2010b ; Morton et al., 2007 ; Vasquéz et al., 2012 ). With this method of extraction, Morton et al.…”

“…A study by Vasquéz et al. ( 2012 ) used 28 epididymides, but the success rate was not stated. Mamani‐Mango et al.…”

Test hipoosmótico en espermatozoides de alpaca (Vicugna pacos) recuperados del conducto deferente

“…1: b-c) al porcentaje de células que muestran enrollamiento leve y "reacción total" al porcentaje de espermatozoides con algún tipo de enrollamiento (RF+RD) que es importante al evaluar protocolos y recomendar uno ideal que es materia de esta investigación (Fig.1) (Jeyendran et al, 1984). Vásquez et al (2012) empleo los términos de reacción fuerte y reacción total tomando en cuenta ambos conceptos para su evaluación, ya que la reacción fuerte está referida al porcentaje de células que La motilidad y concentración se evaluó en el ISAS® utilizando muestras de 5 µl que se analizaron con una lente de 10X en contraste de fase negativo; para la vitalidad se empleó la tinción de Hancock evaluándose con una lente de 40X en el módulo de vitalidad del ISAS®; para la morfometría espermática, se tiñeron éstos con Hemacolor, se evaluó empleando la lente de 100X con aceite de inmersión. Figura 1.…”