Ovarian morphology and internal vis-à-vis non internal laying in relation to triacylglycerol, hormones and their receptors concentration around the age of sexual maturity in broiler breeder hens.

1. Ovarian morphology, serum hormone concentrations of 17-ß-estradiol, progesterone, testosterone, tri-iodothyronine (T3), thyroxine (T4) and triacylglycerol (TAG) were investigated at 23 and 26 weeks of age in broiler breeder hens provided with ad libitum access to feed. Progesterone, oestrogen-ß, thyroid-α and -ß receptor mRNAs were also quantified in the infundibulum at the same ages. 2. A large variation in the ovarian morphology was observed at 23 weeks of age including hens with undeveloped ovaries, non-laying hens with post ovulatory follicles (POF) and a predominance of non-laying hens without a POF. 3. Serum concentrations of triglyceride, 17-ß-estradiol and progesterone at 23 weeks of age were lower in hens with an undeveloped ovary compared with other groups of hens, whereas testosterone, triiodothyronine and thyroxin were higher. 4. At 26 weeks of age, the average number of hierarchical yellow follicles in normal layers was 7.64 ± 0·41 whereas in internal layers, the follicular numbers were significantly greater at 8.66 ± 0·53. The higher follicular numbers in internal layers were associated with higher serum triglyceride and progesterone concentrations. 5. Oestrogen receptor-ß and thyroid receptor-ß mRNA was up regulated in the infundibulum of internal layers compared with normal laying hens at 26 weeks of age.

The role of the male cloacal gland in reproductive success in Japanese quail (Coturnix japonica).

The adult male Japanese quail has a well developed cloacal gland that produces meringue-like white foam. The physiological significance of the cloacal gland and its foam is still unclear. Therefore, we conducted two experiments to establish the functional role of the cloacal gland and its foam in natural mating and oviducal sperm transport. In the first experiment, artificial insemination of equal numbers of spermatozoa diluted in foam extract and normal saline once in a week were used to determine the role of foam in sperm transport in the female oviduct. After artificial insemination, eggs were collected to measure fertility, the duration of the fertile period, sperm holes and attached spermatozoa in the perivitelline membrane. Higher (P<0.05) fertility and greater duration of the fertile period were observed when semen was inseminated along with foam extract compared with normal saline. Further, the sperm holes and trapped spermatozoa in the perivitelline membrane were also higher (P<0.05) in the presence of foam extract. In the second experiment, two males with bigger and smaller cloacal gland areas were allowed to mate with a female. The mating attempts of males with larger cloacal gland were more successful (P<0.05) than males with smaller cloacal glands. Our results indicated that cloacal foam improves sperm transport in the female oviduct and that males with larger cloacal gland areas are preferred during mating.

Influence of chicken native breeds on some physical and biochemical characteristics and short-term storage of semen.

1. The major objective of this study was to examine the influence of 24-h storage of semen at low temperature on semen characteristics and fertilising ability of spermatozoa in two native breeds (Kadaknath-KN, Aseel Peela-AP) and White Leghorn (WL) chicken. 2. Various physical and biochemical properties of freshly ejaculated semen of KN and AP were investigated. Fertility was examined in freshly-ejaculated as well as 24-h-stored (3°C) semen diluted (1:3) with Beltsville Poultry Semen Extender. 3. No significant difference was observed in sperm motility among the different breeds whereas live counts were higher in WL than the native breeds. Body weight, semen volume and sperm concentration were highest in AP, followed by KN and WL. A similar trend was observed in the percentage of dead and morphologically-abnormal spermatozoa. 4. The activity of acid and alkaline phosphatase in seminal plasma were higher in WL than KN, whereas the opposite trend was recorded for glutamic oxaloacetic and pyruvic transaminases. The cholesterol content of semen was highest in AP, followed by KN and WL. Cholesterol was much lower in seminal plasma compared with whole semen but there were no differences between breeds. Mean values of the methylene blue reduction time test were higher in WL than in the native breeds. 5. Fertility and hatchability, using freshly-diluted semen, were poorer in the native breeds than in WL. The pattern of fertility deteriorated further, especially in native fowls, when the birds were inseminated with 24-h-stored semen. 6. In conclusion, variation in physical and biochemical characteristics of semen in native breeds compared to WL correlated with poor fertility after short-term storage of semen.

Role of nitric oxide in ovarian follicular development and egg production in Japanese quail (Coturnix coturnix japonica).

Role of nitric oxide (NO) in regulating the reproductive functions at hypothalamo-hypophysealovarian axis in Japanese quail was studied. In first experiment, metabolites of NO, i.e. nitrite and nitrate (NO2 and NO3) were estimated together in hypothalamus, serum and ovarian follicles of good and poor layers. In the second experiment, different NO modulators such as L-arginine (L-Arg), sodium nitroprusside (SNP) and N(G)-nitro-L-arginine methyl ester, HCl (L-NAME) were administered to the birds. In the first experiment, significantly higher (P < 0.01) NO2 and NO3 levels in serum, hypothalamus and largest (F1) ovarian follicles were observed in good layers as compared to poor layers. Higher (P < 0.05) NO2 and NO3 concentration was observed in F1 follicles than smaller follicles (F2) only in good layers. The NO2 and NO3 concentration was significantly reduced (P < 0.05) in post ovulatory follicles (POFs) in comparison to F1 and F2 follicles. In the second experiment, the serum NO2 and NO3 concentrations were higher (P < 0.05) in the SNP, lower (P < 0.05) in the L-Name group and unchanged in the L-Arg treated group in comparison to control group. compared to control, L-Arg and SNP increased (P < 0.05) the hypothalamic NO2 and NO3 concentration where as L-NAME reduced (P < 0.05) these levels. The NO2 and NO3 concentration was increased (P < 0.05) as the follicle size increased and it was significantly reduced (P < 0.05) in POFs. The higher (P < 0.05) follicular NO2 and NO3 concentration was observed in L-Arg group in comparison to control group. Egg production was also found to be higher (P < 0.05) in L-Arg group whereas it was not different (P > 0.05) in SNP and L-NAME treated groups. The yolk weight and yolk to albumin ratio was reduced (P < 0.05) in L-NAME group in comparison to control group. It may be concluded from the present study that NO plays a key role in regulating follicular development, ovulatory mechanisms and egg production in Japanese quail.

Induction of ovulation in vitro in the hen: dependency of the response to LH on age and rate of lay.

Two experiments were conducted on White Leghorn hens to investigate the effect of different concentrations of LH on the frequency of induction of ovulation in vitro of ovarian follicles collected 18-20 h before the expected time of ovulation from birds of three different ages (7, 18 and 30 months) or of the same age but laying short (three to four) or long (more than seven) sequences of eggs. The ability of LH to induce ovulation was directly related to the age of the donors of the follicles. Follicles derived from hens laying short sequences of eggs were less responsive to the ovulation-inducing effects of LH than were follicles from birds laying long sequences. These observations suggest that the sensitivity of ovarian follicles to the ovulation-inducing effects of LH declines with age and is greater in hens laying long sequences of eggs than those laying short sequences.

Ovarian maturity as modified by partial sinistral ovariectomy and its transplantation in immature domestic fowl.

In laparotomized hens (70d old), about one-fourth part of the left ovary was removed in Group I and about one-half of it in Group II. In Group III, about one-half of the ovary was left intact and the other half was transplanted to the lateral side, near the caudal lobe of the left kidney. In Group IV, the left ovary was removed completely and about one-half of it was transplanted in the same hen as in Group III, and finally, Group V was run as a shamoperated control. On slaughtering, the gonadal status was examined at the age of 130d. The presence of yellow developing follicles were found in all the hens in Group IV (5 5 (b )), followed by Group II (7 9 (b )), Group III at transplanted site (3 5 (ab )), Group I (6 10 (ab )) Group V (1 12 (a )) and Group III at original site (0 5 (a )). The average number of yellow follicles were recorded 4.5 in Group 1, 5.2 in Group II, nil at original site and 6.3 at transplanted site in Group III, 7.6 in Group IV and 3.0 in Group V, considering those hens bearing yellow follicles only. These observations clearly indicated that the shortening of the ovary advances the maturity of the remaining intact part of the ovary. This may be due to more availability of gonadotropins per unit of ovarian tissue. Transplantation of the ovary with severed nerves hastened its development, indicating that some suppressive role on gonadal maturity may be imposed by the intact nerves.

Variation in angiotensin-converting enzyme activity in the testes of prepubertal, pubertal and adult cockerels.

The variation in the level of angiotensin-converting enzyme in the testes of prepubertal, pubertal and adult cockerels were investigated. Maximum levels of angiotensin-converting enzyme were observed at 8 wk of age, followed by a progressive decrease (P/0.01) until 14 wk of age. After this stage, however, a rapid growth rate of the testes and comb was noted. The concentration of angiotensin-converting enzyme in pubertal and adult birds remained at a significantly lower level than in the prepubertal birds. The level of activity of this enzyme was significantly but negatively correlated with age and with the weights of the testes and comb. The high level of this enzyme found in prepubertal cockerels suggests it may be more highly involved in the early rather than later stages of spermatogenesis.

The influence of prepubertal partial sinistral ovariectomy on the subsequent reproductive function of domestic hens (Gallus domesticus ).

Each of 20 White Leghorn hens of 13 to 14 weeks were subjected to partial sinistral ovariectomy and sham-operations. In half of the hens from each group, the percentage of egg production and clutch size were noted until 50 weeks of age. The growing pattern of normal ovarian follicles was also recorded at 26 weeks of age in a rest half ofthe hens in the two groups. The percentage of egg production and the mean and variance of clutch size did not differ significantly (P / 0.05) between the partially ovariectomized and sham-operated groups. The growing yellow follicles (>8 mm) in the rapidly developing phase in these two groups did not vary, although the smaller follicles (4 to 8 mm in diameter) remained significantly (P / 0.01) more in the shamoperated control group than in the partially ovariectomized group. This observation indicates that smaller follicles (4 to 8 mm) developed in the larger (>8 mm) follicles more efficiently in partially ovariectomized hens than in the sham-operated (control) hens. In a second experiment, one group of hens had all the yellow follicles (>8 mm) removed, while a second group of hens was left untreated. On the 3rd and 6th day post treatment, the hens were examined for the presence of ovarian follicles. No significant (P / 0.05) difference in the growing pattern of subsequent follicles (2 to 4 or 4 to 8 mm) was detected due to treatment. These data demonstrate that the mechanisms that regulate follicular growth and atresia are adjust to maintain normal ovulation following partial ovariectomy.

Effects of cold shock treatment on angiotensin-converting enzyme activity and on semen characteristics in roosters and bulls.

Angiotensin-converting enzyme (ACE) activity has been determined in the semen of certain avian and mammalian species as well as its release during cold shock. The maximum and minimum levels of this enzyme were found in mammalian spermatozoa and in seminal plasma, respectively. It was found that ACE activity in mammalian spermatozoa was more pronounced than in the seminal plasma, whereas in the avian species a revers pattern was observed. However, there were no significant differences in ACE activity in spermatozoa and seminal plasma between layer and broiler strains of avian species. By contrast, ACE activity in the spermatozoa and seminal plasma of buffalo bulls was significantly higher (P/ 0.01) than in cattle bulls. Cold shock did not significantly alter semen characteristics in avian species, while a significant (P/ 0.01) decrease in sperm live counts and motility as well as a corresponding increase in morphological abnormalities were observed in the spermatozoa of cattle and buffalo bulls due to cold shock.

Angiotensin converting enzyme activity in female reproductive tract of domestic fowl (Gallus domesticus) and its interactions with semen.

Presence of angiotensin converting enzyme (ACE, EC 3.4.15.1) in female reproductive tract and its interaction with semen has been investigated in domestic fowl. Higher activity of ACE was detected in homogenates of infundibulum region followed by magnum and relatively low level was noticed in rest of the parts of the oviduct (i.e. isthmus, uterus, uterovaginal-junction and vagina). This decreasing order in ACE activity was, however, reversed when these segments were incubated with semen. ACE in the ovarian follicular wall increased with corresponding increase in the follicle size. Presence of some stimulatory or inhibitory substances for ACE in the oviduct of fowl is suggested.

Effects of cadmium salt on phosphomonoesterases activity and fertilizing ability of fowl spermatozoa.

Daily administration of cadmium salt for 25 days (2.5 mg per Kg body weight) in the male domestic fowl caused the end of treatment period. An increased incidences of concentration. Fertility dropped to zero at the end of the treatment period. Activity of acid and alkaline phosphomonoesterases were also drastically reduced by the end of treatment period. An increased incidences of morphological abnormalities of spermatozoa were noticed in the treated birds. After 46 days cessation of the treatment, full recovery of the above measures was found. These alterations suggest the reversible type of effect of cadmium chloride on the spermatozoa of male domestic fowl.

Cloacal gland foam enhances motility and disaggregation of spermatozoa in Japanese quail (Coturnix japonica).

The adult male Japanese quail produces white foam from the cloacal gland, which is transferred to the female proctodeum during natural mating. The physiological role of foam on quail spermatozoa is still unclear. Therefore, attempts have been made to understand the effect of cloacal foam on motility and metabolism of quail spermatozoa. The profile of various biochemical constitutes in the foam extract was investigated. The addition of foam extract to neat semen completely disaggregated the clumps of spermatozoa leading to vigorous motility. The metabolic rate (MBRT) of the spermatozoa was significantly increased with the addition of foam extract. The foam extract was sub fractionated into seven different fractions by using the molecular cut off devices. Among all the seven sub-fractions from the foam extract, the addition of < 1 KDa sub-fraction contained lactate and has enhanced sperm motility and metabolism. Another fraction (3-10 KDa) has non-protein and non-heparin components which completely disaggregated the clumped quail spermatozoa. However, the remaining fractions did not show any effect on quail spermatozoa. It can be concluded from the present investigation that the lactate present in foam might be a fuel for sperm metabolism and motility. Furthermore, low molecular weight (3-10 KDa) components in the foam may responsible for sperm disaggregation.

Characterization of lactate dehydrogenase enzyme in seminal plasma of Japanese quail (Coturnix coturnix japonica).

Lactate dehydrogenase enzyme present in quail seminal plasma has been characterized. Polyacrylamide gel electrophoresis and subsequently with LDH specific staining of seminal plasma revealed a single isozyme in quail semen. Studies on substrate inhibition, pH for optimum activity and inhibitor (urea) indicated the isozyme present in the quail semen has catalytic properties like LDH-1 viz. H-type. Furthermore, unlike other mammalian species, electrophoretic and kinetic investigations did not support the existence of semen specific LDH-X isozyme in quail semen. The effect of exogenous lactate and pyruvate on sperm metabolic activity was also studied. The addition of 1 mM lactate or pyruvate to quail semen increased sperm metabolic activity. Our results suggested that both pyruvate and lactate could be used by quail spermatozoa to maintain their basic functions. Since the H-type isozyme is important for conversion of lactate to pyruvate under anaerobic conditions it was postulated that exogenous lactate being converted into pyruvate via LDH present in semen may be used by sperm mitochondria to generate ATP. During conversion of lactate to pyruvate NADH is being generated that may be useful for maintaining sperm mitochondrial membrane potential.

Fertility and hatchability in RIR and WL breeds as functionally modified by crossing them in alternate sex combinations (Gallus domesticus).

(1) Four breeding groups of Rhode Island Red and White Leghorn domestic fowl (RIR (female) x RIR (male), RIR (female) x WL (male), WL (female) x RIR (male) and WL (female) x WL (male)) were compared for fertility, hatchability, and their post-insemination sustainability, egg weight loss during incubation and uncovered yolk in abdominal cavity of dead in shell in order to understand the problems associated with the RIR breed in these respects. (2) Crossing RIR (female) with WL (male) or in reverse sex combinations did not improve fertility in comparison to pure RIR chickens and all these groups were less fertile than the pure WL. (3) Unlike fertility, hatchability in RIR improved with the change to either sex partner of the WL breed but the WL (female) x RIR (male) combination was similar to the pure WL (97.72 and 97.12%, respectively). In contrast, crossing RIR (female) with WL (male) resulted in an improvement (86.67%) as compared to pure RIR (76.67%) but still lower than the pure WL and WL (female) x RIR (male) cross. (4) Egg weight loss during incubation was more (20.16%) in pure RIR as compared to RIR (female) x WL (male) (17.13%), followed by WL (female) x RIR (male) (10.28%) and pure WL (9.57%). (5) There were more dead-in-shell embryos with yolks outside their abdominal cavity in pure RIR and their crosses as compared to pure WL breeds. (6) Fertility was sustained for longer in WL than other combinations with post-artificial insemination using constant number of spermatozoa. Fertility after a week of insemination tended to decrease more rapidly than hatchability on a fertile egg basis. (7) It is concluded that both sexes are responsible for the poor fertility in RIR but the female is responsible for poor hatchability and this poor performance is mainly due to greater egg weight loss during incubation.

Catecholamines in ovarian follicles during the ovulatory cycle of White Leghorn hens.

Dopamine (DA) concentrations in the largest ovarian follicle (F1) showed significant decrease at 12 and 8 h prior to ovulation. Nor-epinephrine (NE) and epinephrine (E) concentrations in the F1, sampled at different times of the ovulatory cycle, were found to increase with the approach of ovulation time. However, the concentration of epinephrine in the F1 was low just after oviposition compared with the immediate two previous stages investigated. The concentrations of DA, NE and E in the second largest follicles (F2) did not vary significantly during the ovulatory cycle. Average concentration of DA was low and that of NE and E high in the F1 compared with the respective concentrations in the F2.

Effects of cyproterone acetate and testosterone treatments on some physical parameters, angiotensin converting enzyme activity and fertilizing ability of spermatozoa of domestic cocks (Gallus domesticus).

Healthy adult cocks were administered through the intramuscular route with 1 or 15 mg cyproterone acetate (CyA)/kg b. wt. or testosterone at the rate of 1.5 mg/kg b. wt. daily for a period of 25 days. High doses of CyA and testosterone caused severe suppression (P less than 0.01) of semen volume, sperm concentration, sperm motility and angiotensin converting enzyme activity (ACE EC 3.4.15.1). Fertilizing ability of spermatozoa was also reduced drastically at the end of treatment period. No loss of sexual behaviour was observed in treated birds as compared to controls. Recovery in all parameters from deleterious effects of these drugs was rapid after cessation of treatment. Lower doses of CyA did not affect the parameters of semen. The results are discussed in relation to the possible mechanism of action of CyA and testosterone.

Histamine in egg yolk: heat resistance and relation to production status.

1. The presence of histamine in egg yolk, its losses during storage, incubation and heating, and its relationship to cage density stress and the egg production status of hens are described. 2. A significant (P less than 0.05) reduction in the histamine concentration of egg yolk was noted after incubating or heating egg yolks or on storage eggs. 3. Histamine concentration was greater in the yolks of high egg producing hens. 4. The concentrations of histamine in egg yolks did not change significantly (P less than 0.05) on keeping the hens individually in a cage, pairing them in the same size cage or after separating them again to individual cages.