Evaluation of Porcine Sperm Activity Using an Electrochemical Device

[Introduction] In recent years, in vitro fertilization has become an important method in the field of infertility treatment. Research is progressing in the livestock industry to improve fertility and pregnancy rates. The motility of sperm can be evaluated through time-lapse microscopy. This method is cost-effective and requires little space for activity evaluation. In addition, methods for selecting motile sperm include those that utilize centrifugation, such as density gradient centrifugation. However, there is a concern that centrifugation may physically damage sperm due to the large gravitational force it receives. We previously measured the respiratory activity levels of sperm and somatic cells in raw cow milk using amperometry; however, we did not evaluate sperm motility [1]. Therefore, in this study, we developed an electrochemical device that collects motile spermatozoa using moving gravity sedimentation without adding centrifugation, and we reported the examined changes in the oxygen reduction current before and after sperm movement. [Experiment] Porcine (Duroc) sperm (approximately 108 sperm/mL) was purchased from the Miyagi Prefecture Livestock Experiment Station for use. The device was filled with 5.7 mL of 11.4 mM glucose-containing phosphate buffer solution (pH = 7.4), and a three-electrode system was constructed. A platinum electrode (diameter = 1.6 mm) was installed in the middle tube and a platinum electrode was installed in the counter electrode (Fig. 1). The applied voltage was maintained at -0.5 V vs. Ag/AgCl, and the sampling time was 0.1 s. After the start of measurement, 300 µL of the sperm sample was dropped onto the bottom of the outer tube, and amperometry was performed for approximately 60 minutes. [Results and discussion] Fig. 2 shows the amperogram from 40 to 55 minutes after dropping the sperm sample. After approximately 42 minutes, the oxygen reduction current started to decrease. After approximately 10 minutes, the current became constant, and a decrease of approximately 100 nA was observed. This result suggested that the pig sperm potentially consumed oxygen during movement from the bottom of the outer ring to the center of the well. The number of sperm that swam upward and moved to the center was approximately 4.4 × 106 sperm/mL in 60 minutes, and the recovery rate was approximately 15.0%. Compared to the literature value for human sperm (approximately 13.8%), the experimental results were considered reasonable, suggesting the possibility of capturing the real-time respiratory activity of moving sperm [2]. In this paper, we thoroughly discussed the relationship between the initiation of porcine sperm migration and the oxygen reduction current. [References] [1]S.Kasai, A.Prasad, R.Kumagai, K.Takanohashi, Biology,2022,11,549.https://doi.org/10.3390/biology11040549. [2] K.Tatsumi, T.Tatsumi, T.Uchida, K.Saito, H.Saito, F&S Reports, 2020,1(2), 106-112. Figure 1