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The acrosome reaction (AR) is a strictly-regulated, synchronous exocytosis that is required for sperm to penetrate ova. This all-or-nothing process occurs only once in the sperm lifecycle through a sequence of signaling pathways. Spontaneous, premature AR therefore compromises fertilization potential. Although protein kinase A (PKA) pathways play a central role in AR across species, the signaling network used for AR induction is poorly understood in birds. Mechanistic studies of mammalian sperm AR demonstrate that PKA activity is downstreamly regulated by Src family kinases (SFKs). Using SFK inhibitors, our study shows that in chicken sperm, SFKs play a role in the regulation of PKA activity and spontaneous AR without affecting motility. Furthermore, we examined the nature of SFK phosphorylation using PKA and protein tyrosine phosphatase inhibitors, which demonstrated that unlike in mammals, SFK phosphorylation in birds does not occur downstream of PKA and is primarily regulated by calcium-dependent tyrosine phosphatase activity. Functional characterization of SFKs in chicken sperm showed that SFK activation modulates the membrane potential and plays a role in inhibiting spontaneous AR. Employing biochemical isolation, we also found that membrane rafts are involved in the regulation of SFK phosphorylation. This study demonstrates a unique mechanism for regulating AR induction inherent to avian sperm that ensure fertilization potential despite prolonged storage.

Glucose plays an important role in sperm flagellar motility and fertility via glycolysis and oxidative phosphorylation, although the primary mechanisms for ATP generation vary between species. The glucose transporter 1 (GLUT1) is a high-affinity isoform and a major glucose transporter in mammalian spermatozoa. However, in avian spermatozoa, the glucose metabolic pathways are poorly characterised. This study demonstrates that GLUT1 plays a major role in glucose-mediated motility of chicken spermatozoa. Using specific antibodies and ligand, we found that GLUT1 was specifically localised to the midpiece. Sperm motility analysis showed that glucose supported sperm movement during incubation for 0–80min. However, this was abolished by the addition of a GLUT1 inhibitor, concomitant with a substantial decrease in glucose uptake and ATP production, followed by elevated mitochondrial activity in response to glucose addition. More potent inhibition of ATP production and mitochondrial activity was observed in response to treatment with uncouplers of oxidative phosphorylation. Because mitochondrial inhibition only reduced a subset of sperm movements, we investigated the localisation of the glycolytic pathway and showed glyceraldehyde-3-phosphate dehydrogenase and hexokinase I at the midpiece and principal piece of the flagellum. The results of this study provide new insights into the mechanisms involved in ATP production pathways in avian spermatozoa.

Pengembangan Domba Lokal melalui program Inseminasi Buatan memerlukan informasi mengenai kualitas semen, khususnya semen beku yang dalam pembuatannya memerlukan gliserol sebagai krioprotektan dengan kadar yang tepat. Penelitian ini bertujuan untuk mengetahui kualitas semen Domba Lokal dan mengetahui level gliserol yang menghasilkan kualitas semen post-thawing yang paling optimal. Penelitian dilakukan dengan menggunakan rancangan acak lengkap (RAL) dengan empat perlakuan level gliserol (4%, 5%, 6% dan 7%) dan 14 kali ulangan. Parameter yang diamati meliputi motilitas, abnormalitas dan recovery rate. Data yang diperoleh dianalisis menggunakan analisis ragam (ANAVA) dan perbedaan antar perlakuan diuji menggunakan Uji lanjut Duncan. Hasil penelitian menunjukkan bahwa level gliserol berpengaruh nyata (p<0.05) terhadap motilitas dan recovery rate namun tidak berpengaruh nyata terhadap abnormalitas sperma. Perlakuan level gliserol 5% nyata (p<0.05) menghasilkan motilitas (40,47%) dan recovery rate (46,53%) tertinggi dibanding perlakuan lainnya. Disimpulkan bahwa semen Domba Lokal memiliki kualitas yang baik dan memenuhi syarat untuk dilakukan pengolahan lebih lanjut sampai semen beku dengan level gliserol 5% menghasilkan kualitas semen post-thawing yang paling optimal.