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Simple Summary Progesterone is an endogenous steroid hormone, which can induce capacitation and/or acrosome reactions in semen of certain mammalian species. Our study aimed to investigate the effect of progesterone on the functional status of fresh bovine spermatozoa using a chlortetracycline fluorescent probe. Results showed that heparin induced capacitation in spermatozoa incubated with or without progesterone. The destruction of microfilaments by an inhibitor of cytochalasin D blocked the stimulating effect of heparin. Steroid hormone in mixture with prolactin stimulated the acrosome reaction in spermatozoa, which was blocked by an inhibitor of microtubule polymerization (nocodazole). At the acrosome stage, prolactin provided the undergoing of acrosome reaction in male gametes. This effect was noted both in the presence and absence of progesterone and inhibited by nocodazole. The supplementation of dibutyryl cyclic adenosine monophosphate during the acrosome reaction to progesterone-untreated spermatozoa did not cause changes in proportion of acrosome-reacted cells. However, when progesterone was added during capacitation, a significant increase in the proportion of capacitated cells was noted, which was inhibited by nocodazole. Thus, progesterone under the action of prolactin and dibutyryl cyclic adenosine monophosphate determines the functional status of fresh spermatozoa, which indicates progesterone-modulating effect on the indicators of post-ejaculatory maturation of male gametes. Abstract The aim of this study is to identify the effects of progesterone (PRG) on the capacitation and the acrosome reaction in bovine spermatozoa. The fresh sperm samples were incubated with and without capacitation inductors (heparin, dibutyryl cyclic adenosine monophosphate (dbcAMP)), hormones (prolactin (PRL), PRG), inhibitors of microfilaments (cytochalasin D) and microtubules (nocodazole) during capacitation and acrosome reactions. The functional status of spermatozoa was examined using the chlortetracycline assay. Supplementation of heparin stimulated capacitation in the presence and absence of PRG. Cytochalasin D blocked the stimulating effect of heparin on capacitation. The addition of PRL during capacitation (without PRG) did not affect the functional status of spermatozoa, while in PRG-treated cells PRL stimulated the acrosome reaction. PRL (with and without PRG) increased the acrosome reaction in capacitated cells. These PRL-dependent effects were inhibited by nocodazole. During the acrosome reaction, in presence of dbcAMP, PRG decreased the proportion of acrosome-reacted cells compared to PRG-untreated cells. This effect in PRG-treated cells was canceled in the presence of nocodazole. In conclusion, PRG under the action of PRL and dbcAMP determines the changes in the functional status of native sperm cells, which indicates PRG modulating effect on the indicators of post-ejaculatory maturation of spermatozoa.

Spermatogonia are early-undifferentiated germ cells, giving rise to mature male generative cells — the spermatozoa. There are two types of spermatogonia – A and B. Of greatest interest is the use of type A spermatogonia, which are the stem cells of the testes. To select the appropriate age for collecting spermatogonia А from quails it is necessary to know the specific features of spermatogenesis. Development dynamics of various spermatogonia types in the quail testicular tubules was studied. Histological studies of the quails testicular tubules at the age of 1, 2, 3, 4, 5 and 6 weeks (n = 30) were carried out. Samples of testis tissue were fixed in Bouin’s fixative. Histological sections were stained with hematoxylin-eosin. Identification of different spermatogonia types was carried out according to their morphology. Type A spermatogonia were additionally identified by immunohistochemistry using SSEA-1 antibodies. The proportion of spermatogonia in the total number of spermatogenic cells in the seminiferous tubules of quails changed with age. The maximum value was reached at the age of 3 weeks and it was 76±6%. On reaching maturity (6 weeks), this indicator decreased to 12 ± 1 %. In the early period of ontogenesis (1–2 weeks), spermatogonia cells were represented mainly by type A spermatogonia. The proportion of these cells from the total number of spermatogonia reached 80 ± 3 %. With increasing age, this indicator decreased, reaching minimum values for achieving maturity (6 weeks) - 16 ± 1 %. The percentage of type B spermatogonia in the seminiferous tubule of quails on the contrary increased with age — from 5 ± 1% at 1 week old to 70 ± 2% at maturity. Thus, the age no later than 2 weeks is the most optimal for the isolation type A spermatogonia of quails. Supported by RFBR (18-29-07079).

The cells of the male gonads are considered as a valuable genetic material for the conservation of the gene pool of breeds and lines of agricultural birds, as well as the directed modification of the poultry genome. Mature germ cells – spermatozoa and their predecessors – spermatogonia, spermatocytes and spermatids can be used for these purposes. To obtain these types of cells, it is necessary to know the characteristics of their development (spermatogenesis). The dynamics of the development of certain spermatogenic cell types in the testicular tubules of different-aged roosters has been studied. Histological studies were performed on testes of roosters aged from 1 week to 6 months with an interval of 2 weeks. Samples of testis tissue were fixed in Bouin’s solution. Histological sections were stained with hematoxylin-eosin. Identification of different cell types (Sertoli, spermatogonia, spermatocytes, spermatids, sperm cells) was carried out according to their morphology. At the age of 1–6 weeks in the seminiferous tubule of roosters, the mainly presence of two cell types was noted: Sertoli cells and spermatogonia. From 7 weeks of age, spermatocytes were detected in the seminiferous tubules, in the 4 months - spermatids, in the 5.5 months - sperm cells. The number of Sertoli cells remained almost unchanged with age and was 21 ± 2. The percentage of these cells decreased with age from 71 ± 3 % to 5 ± 1 %. The percentage of spermatogonia also decreased with age from 75 ± 2 % to 7 ± 1 %. The number of spermatids and spermatozoa, on the contrary, increased to puberty (6 months) and reached 54 %. The study was supported by the RFBR within Project no.18-29-07079.

Cryopreservation of testicular stem cells - spermatogonia is of interest along with the creation of semen cryobanks. During transplantation into recipients’ testes, spermatogenic cells can create a significant population of germ cells in the process of differentiation. The knowledge about spermatogenesis course in males is necessary for the effective selection spermatogenic cells. The research aim was to study the age-related characteristics of spermatogenesis in geese. The histostructure of gander testes (n = 35) at the age of 1 to 7 months was studied. The diameter of seminiferous tubules, and the types and number of spermatogenic cells in them were evaluated. From each gander at least 30 seminiferous tubules were examined. At the age of 1 month, the diameter of the seminiferous tubules was 51±1 μm. In subsequent age periods, this indicator increased and amounted to 63±2, 65±3, 66±2, 79±3, 98±6 and 170±5 μm at the age of 2, 3, 4, 5, 6 and 7 months, respectively. Diameter increase with the age was associated with an increase of spermatogenic cells number inside tubules. At the 1 months age, the number of spermatogenic cells in one seminiferous tubule did not exceed 22±1. At the age of 2, 3, 4, 5, 6, and 7 months, this indicator increased by 2.1, 2.7, 3.1, 6.4, 8.5 and 21.2 times. At the age from 1 to 3 months, the main cells types were Sertoli and spermatogonia cells. Primary and secondary spermatocytes from 4 months of age and spermatids from 5 months of age were visualized in the seminiferous tubules. Sperm were detected in the seminiferous tubules at 6 months old, the number of which increased towards the age of 7 months. The study was supported by RSF within Project №16–16–04104.

The use of testicular stem cells (spermatogonia) is of most interest for obtaining individuals with predetermined traits and genome genetic modification and for conservation of poultry gene pool. A significant population of mature donor germ cells (sperm) is formed upon successful spermatogonia cells transplantation into the testes of male recipients. Obtained sperm can be used to produce offspring with the desired traits. A key step in this technology is the removal of own spermatogenic cells (inhibition of spermatogenesis) in male recipients. The aim of research was to develop and optimize methodological approaches to inhibit the spermatogenesis in quail using busulfan. This drug was injected directly into the testes parenchyma of mature males by multiple injection at the concentration from 20 to 100 mg per 1kg of body weight (n = 25). Histological preparations of testes from the experimental quails were obtained to study composition of spermatogenic cells in the seminiferous tubules after busulfan administration. The male peers who were not injected with busulfan were used as a control. Experimental quails showed a decrease in the number of spermatogenic cells in the seminiferous tubules 32, 75, 111, 119 and 118 times compared with the control when using busulfan in concentrations 20, 40, 60, 80 and 100 mg/kg of weight, respectively (P < 0.001). The cells composition in the seminiferous tubules from experimental quails was represented mainly by Sertoli cells and spermatogonia. After busulfan introduction at the concentrations 20, 40, 60, 80 and 100 mg/kg, the percentage of spermatogonia was 55±5 %, 24±4 %, 6±2 %, 5±2 % and 4±1 %, respectively. The use of busulfan at the concentration of 80–100 mg/kg led to high mortality of quails. Thus, it was found that the optimal busulfan concentration for elimination of quail spermatogenic cells was 60 mg/kg. Supported by RFBR within Project №18-29-07079.