V. Yu. Denisenko, T. I. Kuzmina
All-Russian Research Institute of Genetics and Breeding of Farm Animals, branch of Federal Scientific Center of Animal Husbandry – Ernst All-Russian Research Institute of Animal Husbandry, Moskovskoe sh., 55a, Sankt-Peterburg, 196601, Russian Federation
Abstract. We investigated the participation of the protein kinases A and C in the activation of capacitation processes on bulls’ spermatozoids. Knowing the intracellular mechanisms associated with capacitation will allow affecting spermatozoids to improve fertilization results. Using the CTC test, which determines the location of chlortetracycline fluorescence in spermatozoid, we studied their distribution into groups with different functional status – uncapacitated, capacitated, and acrosome-reactive. Spermatozoids from three bulls were examined; each group was taken at least 200 spermatozoids. The compound Ro 31-8220 (10 ng/mL) was used as a protein kinase C inhibitor; the compound N-89 (10 umol) was used as a protein kinase A inhibitor. We evaluated how substances, which stimulate the release of Ca2+ from intracellular depots and promote capacitation, influence spermatozoids. These substances included dibutyryl cyclic adenosine monophosphate (dbcAMP, 100 umol) and guanosine triphosphate (GTP, 10 umol) in the presence of progesterone (1 ug/mL). The number of capacitated spermatozoid with dbcAMP + GTP in the variants without inhibitors and with the protein kinase C inhibitor did not have significant differences – (47.3 ± 2.4)% and (42.5 ± 1.6)%, respectively. The protein kinase A inhibitor significantly reduced the number of capacitated spermatozoid cells during incubation with dbcAMP and GTP to (23.1 ± 1.8)%, compared with the version without the inhibitor. The fluorescence intensity of intracellular calcium depots was determined using a chlortetracycline fluorescence probe. In the presence of dbcAMP + GTP in variants without inhibitors and with the protein kinase C inhibitor, it did not differ significantly and amounted to (72.3 ± 1.6)% and (70.0 ± 1.9)%, respectively. The protein kinase A inhibitor cancelled the additional release of Ca2+ from the intracellular spermatozoid depot, activated by the combined action of dbcAMP and GTP. The fluorescence intensity of chlortetracycline in the presence of dbcAMP + GTP was significantly higher in the variant with the protein kinase A inhibitor than in the variant without the inhibitor and amounted to (85.0 ± 1.7)% and (72.3 ± 1.6)%, respectively. Consequently, in the case of combined action of dbcAMP and GTP, protein kinase A increases the number of capacitated spermatozoid.
Keywords: protein kinases; capacitation; calcium; intracellular depots; bull spermatozoids.
Author Details: V. Yu. Denisenko, D. Sc. (Biol.), leading research fellow; T. I. Kuzmina, D. Sc. (Biol.), head of laboratory (e-mail: Этот адрес электронной почты защищён от спам-ботов. У вас должен быть включен JavaScript для просмотра.).
For citation: Denisenko VYu, Kuzmina TI. [Features of the mobilization of calcium from intracellular depots during the capacitation of bulls’ spermatozoids]. Dostizheniya nauki i tekhniki APK. 2020;34(2):65-8. Russian. doi: 10.24411/0235-2451-2020-10214.