Summary. Exposure of bovine luteal tissue to the anti-microfilament agent cytochalasin B inhibited both basal and LH-induced steroidogenesis, but had no effect on the portion of the progesterone secreted into the incubation medium. Addition of 10% calf serum to the incubation medium caused no significant increase in progesterone synthesis, but produced a significant increase in the portion of the progesterone found in the incubation medium. This increased secretion in the serum-containing medium was not altered by the presence of LH, and cytochalasin B was unable to prevent it. Exposure of the luteal tissue to the antimicrotubular drugs, colchicine and vinblastine, produced no alteration in synthesis or secretion of progesterone, even when LH was present, and neither drug inhibited the serum-induced increase in secretion of progesterone. These studies suggest that luteal cell microfilaments may be involved in LH-stimulated progesterone production while microtubules apparently play no direct role in progesterone synthesis.
P. J. Battista and W. A. Condon
Summary. Incubation of bovine luteal cells with the alternative pathway catecholamines octopamine, synephrine and deoxyadrenaline at concentrations of 10 −6 to 10−3 m enhanced the production of progesterone (P < 0·05). Tryamine did not alter basal progesterone production (P > 0·05). Addition of noradrenaline and adrenaline at concentrations of 10−4 to 10−7 m significantly elevated the production of progesterone (P < 0·05). The steroidogenic response to noradrenaline and adrenaline was greater than that for octopamine, synephrine and deoxyadrenaline (P < 0·05). Response to both primary (10−6 m) and alternative (10−4 m) pathway catecholamines was inhibited by propranolol (10−5 m, P < 0·05) but not phentolamine (10−5 m, P > 0·05). These results demonstrate that octopamine, synephrine and deoxyadrenaline can affect steroidogenesis by bovine luteal cells, and their action is mediated by β-adrenergic receptors.
P. J. Battista and W. A. Condon
Summary. The addition of acetylcholine or histamine (10− 7 to 10−4 m), γ-aminobutyric acid, a dopamine agonist, and melatonin (10−7 to 10−5 m) did not alter basal or LH-stimulated progesterone production (P>0.05). The addition of the specific β2-adrenergic agonist terbutaline and salbutamol did not significantly elevate progesterone production. Treatment of luteal cells with serotonin (5-HT), 10−6 to 10−4 m, increased the production of progesterone (P<0.05). This stimulated production was inhibited by the addition of mianserin (10−5 m, a 5-HT antagonist; P<0.05). Isoproterenol (10−7 to (10−4 m) also resulted in significant increases in progesterone production (P<0.05). The combined treatments of 5-HT + LH, isoproterenol+ LH, or isoproterenol + 5-HT did not result in a further increase in progesterone above that observed in response to LH or isoproterenol alone (P>0.05). The isoproterenol-induced progesterone production could not be blocked by butoxamine (10−5 m, a β2-antagonist), or practolol (10−5 m, a β1-antagonist), but was inhibited by propranolol (10−5 m, a general β-antagonist; P<0.05). The response to isoproterenol was unaffected by mianserin (10−5 m). These results demonstrate a possible role for 5-HT in the regulation of steroidogenesis by the corpus luteum of the cow. Furthermore, these results suggest that serotonin-induced progesterone production is a receptor-mediated event.
J. L. Pate and W. A. Condon
Summary. Bovine luteal cells can utilize low density lipoprotein (LDL) or high density lipoprotein (HDL) as a source of cholesterol for steroidogenesis, and administration of PGF-2α in vitro suppresses lipoprotein utilization. The objective of this study was to examine the mechanism by which PGF-2α exerts this effect. Cultured bovine luteal cells received 0·25 μCi[14C]acetate/ml, to assess rates of de-novo sterol and steroid synthesis, with or without lipoproteins. Both LDL and HDL enhanced progesterone production (P < 0·01), but caused a significant reduction in the amount of radioactivity in the cholesterol fraction. PGF-2α treatment inhibited the increase in lipoprotein-induced progesterone synthesis (P < 0·01), but did not prevent the reduction in de-novo cholesterol synthesis brought about by LDL or HDL. PGF-2α alone reduced cholesterol synthesis (P < 0·01), but it was not as effective as either LDL or HDL. Both lipoproteins and PGF-2α also decreased the amount of radioactivity in the progesterone fraction (P < 0·01), and the effect of PGF-2α was similar to that of the lipoproteins. It is concluded that lipoproteins can enhance progesterone production and also suppress de-novo cholesterol synthesis in bovine luteal cells, but only the former effect of lipoproteins is inhibited by PGF-2α. Therefore, it is suggested that PGF-2α allows entry of lipoprotein cholesterol into the cell, but prevents utilization for steroidogenesis. In addition, PGF-2α alone can suppress cholesterol synthesis, as well as decrease conversion of cholesterol to progesterone.
Keywords: corpus luteum; PGF-2α; lipoproteins; cholesterol; cow
J. P. Poff, D.L. Fairchild, and W. A. Condon
Summary. Corpora lutea were removed from regularly cycling dairy cows, dissociated with collagenase and cultured for 8 or 10 days in Ham's F-12 medium. In Exp. 1 treatment with insulin, or an insulin–transferrin–selenium combination (ITS), increased progesterone production from basal levels on Day 4 of culture to 234% (P < 0·01) above controls on Day 10. LH alone increased progesterone production 45% above controls on Day 10 (P > 0·05). When LH was combined with insulin or ITS, progesterone production was stimulated to an average of 1802% (P < 0·01) above controls on Day 10 of culture. Transferrin or selenium without insulin did not allow LH to stimulate progesterone synthesis. In Exp. II, LH alone or LH plus gentamicin or penicillin–streptomycin increased progesterone production from basal levels on Day 2 steadily to an average of 468% (P < 0·01) above controls (no antibiotics) by Day 8 of culture. The addition of amphotericin-B, alone or in combination with the other antibiotics, inhibited all LH-stimulated progesterone synthesis, but did not affect basal progesterone levels. We conclude that insulin is essential for maximal steroidogenesis in a bovine luteal cell culture system, and that LH-stimulated progesterone production is inhibited in the presence of amphotericin-B, but is not inhibited by gentamicin or penicillin–streptomycin. The elimination of amphotericin-B, coupled with the addition of insulin to the cell culture system increased the responsiveness of the cells to LH. These culture conditions represent the first report in which LH increased total progesterone production for 10 days, maintaining luteal function in a chemically-defined culture system.
Keywords: antibiotics; luteal cells; progesterone synthesis
P. J. Battista, J. P. Poff, D. R. Deaver, and W. A. Condon
Summary. Biogenic amines were administered using osmotic pumps placed subcutaneously in the neck region of regularly cycling, non-lactating dairy cows on Days 9–11 (oestrus = Day 0) of the oestrous cycle. Blood samples were collected using indwelling jugular catheters and the plasma progesterone concentrations were measured. Samples were collected at 4-h intervals for the first 12 h of treatment and thereafter at 12-h intervals for the remainder of the 72-h treatment period. After administration of various doses of noradrenalne, adrenaline and serotonin (0·5–2·0 μg/kg/h) significant elevation of plasma progesterone was achieved at a dosage of 2·0 μg/kg/h (P < 0·01). The response to adrenaline was greater than that observed for noradrenaline and serotonin (P < 0·05). Within-treatment comparison to pretreatment samples showed plasma progesterone concentrations to increase within 4 h after the administration of noradrenaline, adrenaline and serotonin (P < 0·05) and this enhancement was maintained throughout the treatment period (P < 0·05). The elevation in plasma progesterone concentrations induced by noradrenaline, adrenaline and serotonin was independent of changes in circulating concentrations of luteinizing hormone. These results support a physiological role for endogenous biogenic amines in the control of bovine luteal progesterone production.