Isolation and culture of ovine and bubaline small and large pre-antral follicles: effect of cyclicity and presence of a dominant follicle.

Studies were conducted to examine the effects of the cyclicity and the presence of a dominant follicle (DF) in ovary on the recovery and in vitro growth of pre-antral follicles (PFs) in sheep and buffalo. Small pre-antral follicles (SPFs, 100-250 microm) and large pre-antral follicles (LPFs, 250-450 microm) were isolated from slaughterhouse ovaries in the breeding seasons by a mechanical and enzymatic method. The sheep and buffalo PFs were cultured in vitro for 6 and 15 days, respectively, and examined for their growth, survival and antrum formation rates and growth rates of oocytes in cultured pre-antral follicles. The follicles of the sheep and buffalo were recovered and cultured simultaneously within replicates. The recovery rates (number per ovary) of both SPFs and LPFs were significantly (p < 0.05) higher in cyclic ewes (SPFs: 22.0 +/- 3.3 vs 12.1 +/- 2.6 and LPFs: 16.0 +/- 3.6 vs 9.2 +/- 1.8) and buffaloes (SPFs: 9.2 +/- 1.3 vs 4.1 +/- 1.0 and LPFs: 10.3 +/- 2.7 vs 5.4 +/- 0.7) compared with those recovered from acyclic ones. Presence of a DF in ovary significantly (p < 0.05) reduced the recovery rates of LPFs in ewes (9.06 +/- 2.7 vs 16.4 +/- 3.8) but had no effect in buffalo. Cyclicity of animals or follicular dominance had no effects on in vitro growth, survival and antrum formation rates and growth rates of oocytes in cultured PFs of SPFs and LPFs in both sheep and buffalo. The in vitro growth, survival and antrum formation rates of LPFs and growth rates of oocytes in cultured LPFs were significantly (p < 0.05) higher than those observed in SPFs in both sheep and buffalo. The overall recovery and growth rates of the PFs were lower in buffaloes compared with ewes.

Follicular fluid concentrations of glucose, lactate and pyruvate in buffalo and sheep, and their effects on cultured oocytes, granulosa and cumulus cells.

The objective was to determine ovarian follicular fluid concentrations of glucose, lactate, and pyruvate in relation to follicle size in buffalo and sheep. The effect of varying concentrations of these substances on in vitro oocyte maturation, oocyte protein content, and granulosa and cumulus cell growth was also investigated. Follicular fluid was aspirated from various sizes of follicles (from ovaries without a dominant follicle) collected from adult, cycling nonpregnant buffalo (Bubalus bubalis) and sheep (Ovis aries) during the breeding season. Overall, mean (+/-S.E.M.) concentrations (mM) were glucose 2.42+/-0.31 and 1.40+/-0.22, lactate 7.56+/-2.61 and 10.42+/-1.64, and pyruvate 0.02+/-0.01 and 0.002+/-0.00, in buffalo and sheep, respectively. In both species, as follicles became larger, concentrations of glucose significantly increased, lactate significantly decreased, but pyruvate was not affected. Oocyte maturation was higher (P<0.05) in medium containing supra-physiological concentrations of either glucose (5 mM), or pyruvate (10 mM) alone, or physiological concentrations of glucose, lactate and pyruvate in combination, compared to supra-physiological concentrations of lactate (15 mM) alone, or sub- or supra-physiological concentrations of glucose, lactate and pyruvate in combination (both species). The protein content of oocytes was not significantly affected by the concentration of glucose, lactate, and pyruvate in the maturation medium. However, growth of granulosa and cumulus cells was higher (P<0.05) in medium containing supra-physiological concentrations of glucose (5 mM) alone, or pyruvate (10 mM) alone, or physiological, or supra-physiological concentrations of glucose, lactate and pyruvate in combination, compared to supra-physiological concentrations of lactate (15 mM) alone, or sub-physiological concentrations of glucose, lactate and pyruvate in combination (both species). In conclusion, concentrations of glucose, pyruvate and lactate in the medium had cell type-specific effects on oocyte maturation, and on growth of granulosa and cumulus cells. Furthermore, glucose and pyruvate were the principal energy sources for oocytes and follicular somatic cells in buffalo and sheep.

Effect of alliums on total lipid in plasma and tissues of Japanese quails (Coturnix coturnix japonica) with respect to age and sex.

Dietary onion and garlic caused an increase in the level of liver and plasma total lipids in Coturnix coturnix japonica. This increase could be due to the effect of an increased feed intake, bile production, digestion and absorption that in turn caused an increased utilisation of dietary fat, increased transfer of dietary lipids to the liver and/or due to increased lipogenesis as such. However, there was no increase in the muscle lipid content. This effect in the muscle could be due to inhibitory effect of onion or garlic on lipoprotein lipase activity. Further, the changes in the tissue total lipid level in the control group due to change in age and sex were also observed.

Effect of extracts of Murraya koenigii leaves on the levels of blood glucose and plasma insulin in alloxan-induced diabetic rats.

The effect of daily oral administration of aqueous extract (600 mg/kg b.wt.) and methanol extract (200 mg/kg b.wt.) of Murraya koenigii Spreng leaves for a period of eight weeks was studied on blood glucose and plasma insulin level in alloxan-induced diabetic rats. Blood glucose levels of diabetic rats treated with aqueous and methanol extracts of Murraya koenigii Spreng showed significant reduction (P<0.05) as compared to diabetic control groups. Plasma insulin showed significantly high on 43rd and 58th days of treatment in aqueous and methanol extracts of Murraya koenigii treated groups. This suggests that the hypoglycemic effect may be mediated through stimulating insulin synthesis and/or secretion from the beta cells of pancreatic islets of Langerhans.

Effect of a partially purified 30.1 kDa ovine follicular fluid protein on ovine follicle and ovarian somatic cell growth, and oocyte maturation in vitro.

AIM: Regulation of folliculogenesis and oocyte-somatic cell interactions in the ovarian follicles is under the control of gonadotrophins and various local factors. In the present study, an attempt was made to isolate and examine the biological activities of ovarian follicular fluid protein(s) in sheep in vitro. METHODS: Follicular fluids aspirated from ovarian follicles of slaughterhouse-derived ovaries were made cell free by centrifugation (5000 g for 30 min) and steroid free by charcoal treatment. The follicular fluid was then subjected to ammonium sulphate precipitation and gel filtration chromatography using G-75 Sephadex. Protein detection was performed using a UV spectrophotometer at 280 nm. The 35-50% fraction yielded a detectable peak and a protein of 30.1 kDa as examined by SDS-PAGE. The effect of increasing doses of the 30.1 kDa ovine follicular fluid protein (oFFP) was tested at different doses on pre-antral and antral follicle growth; cumulus cell expansion; oocyte maturation; changes in protein, calcium and phosphorus levels of oocytes after culture in media containing different levels of isolated protein; mural granulosa cell, polar granulosa cell (cumulus cell), oviductal epithelial cell monolayer formation and granulosa cell proliferation in vitro. RESULTS: The oFFP significantly inhibited antral follicle growth, cumulus expansion, oocyte maturation and somatic cell growth in vitro in a dose-dependent manner. The oFFP did not have a significant effect on the pre-antral follicle growth in vitro. The protein, calcium and phosphorus contents of oocytes were found to decrease in oocytes cultured in maturation medium containing the oFFP. CONCLUSION: The present study demonstrates a follicular fluid factor regulating folliculogenesis and oocyte maturation in sheep.