Vasantha Padmanabhan

Estradiol-17β stimulates basal and thyrotropin releasing hormone induced prolactin secretion by bovine pituitary cells in primary culture

Abstract A series of experiments were conducted which demonstrate that estradiol-17β directly affects bovine pituitary cells in primary culture causing an increase in basal and thyrotropin releasing hormone (TRH)-induced prolactin secretion. Prolactin release by pituitary cells incubated with TRH at concentrations of 0.001, 0.01, 0.1 and 1 ng/ml increased linearly with increasing log concentrations. Exposure of pituitary cells to 5, 50 or 500 ng/ml estradiol for 4 h did not affect basal or TRH-induced prolactin release. However, when the period of exposure to estradiol was prolonged to 6, 12, or 24 h, 0.5, 5 or 50 ng estradiol/ml medium caused pituitary cells to release more prolactin and there was more total prolactin in the system (medium +cell content) than for comparable controls. These increases were linearly related to increasing log concentrations of estradiol used. To determine the chronic effect of estradiol on prolactin secretion, pituitary cells were incubated with estradiol-17β for 11 days during which medium was collected at 24 h intervals beginning on day 3. On day 3, prolactin accumulation in medium of control cultures averaged 2.5 ng/ml, and decreased gradually reaching relatively low levels by day 11 (100 ng/ml). Although prolactin secretion decreased during the culture period, stimulatory effects of estradiol were evident throughout. In addition, these cells still released prolactin in response to TRH (1 ng/ml) on day 11 and magnitude of TRH-induced prolactin release increased with increasing concentrations of estradiol-17β. We conclude that estradiol will increase basal and TRH-induced prolactin release by bovine lactotrophs. These results are consistent with the view that the increase in estradiol that occurs at the end of pregnancy in cattle, may participate in the prolactin surge that occurs at parturition in this species.

Modulation of growth hormone-releasing factor-induced release of growth hormone from bovine pituitary cells☆

Growth hormone (GH)-releasing factor (GRF) at concentrations of 10(-12) through 10(-7) M for 6 hr linearly increased GH release (b1 = 10.4 +/- .3) from bovine anterior pituitary cells in culture. Maximum release of GH (262% above controls) occurred at 10(-7) M GRF. In contrast, GH release-inhibiting factor (SRIF) at 10(-12) through 10(-5) M had no effect on basal concentrations of GH. In a second experiment, as the proportion of SRIF relative to GRF increased, SRIF suppression of GRF-induced GH release from anterior pituitary cells increased. In a third experiment, anterior pituitary cells cultured in media containing fetal calf serum (FCS) were treated with cortisol (0 or 10 ng/ml media) for 24 hr before exposure to 10(-13) through 10(-7) M GRF. GRF linearly increased GH secretion (b1 = 7.4 +/- .3) and cortisol augmented this response (b1 = 10.5 +/- .6). However, when cells were cultured in media containing dextran-charcoal treated FCS, cortisol did not alter GRF-induced GH release. Our results demonstrate that GH response of bovine anterior pituitary cells to GRF was modulated negatively by SRIF. However, augmentation of GRF-induced GH release by cortisol was evident only when cells were cultured in media supplemented with untreated FCS.

Does time of exposure to estradiol and LHRH effect LH release from bovine pituitary cells

Summary Time course of 17-β estradiol and luteinizing hormone-releasing hormone effect on LH release was studied using bovine pituitary cells on day 5 of culture. LHRH at concentrations of .1, 1, 10 and 100 ng/ml increased LH in medium linearly with increasing log concentration of LHRH when present for .75, 1.5, 3, 6 and 24 hr and the percent increase over controls was same at each time period. In addition, estradiol (present for 6, 12 or 24 hr) at .5, 5, and 50 ng/ml also increased LH release linearly both in the presence or absence of LHRH. We conclude that the stimulatory effect of LHRH on LH release from bovine pituitary is consistant over 24 hr and the stimulatory effect of E2 on both basal and LHRH induced LH release may be mediaed at least in part directly on the pituitary. The authors acknowledge Dr. R. R. Neitzel and L. T. Chapin for valuable assistance with computer programming and P. Harkins and C. Wallace for technical help.

Pineal Compounds Alter Prolactin Release from Bovine Pituitary Cells

Summary Compounds native to the pineal were screened using a bovine pituitary cell culture system to investigate their effects on pituitary prolactin secretion. Thyrotropin releasing hormone (10−11-10−7 M) and arginine vasotocin (10−10-10−6 M) increased prolactin in medium and this increase was linearly related to increasing log concentrations of each peptide. In contrast, dopamine (10−9-10−7 M) and norepinephrine (10−10-10−8 M) decreased prolactin accumulation in medium and decrease was linearly related to increasing log concentrations of catecholamines. Serotonin (10−10-10−6 M) also inhibited prolactin release and decrease was linearly related to increasing log concentrations of serotonin. Melatonin, 5-hydroxytryptophol and 5-methoxytryptophol had no direct effect on pituitary prolactin secretion. We conclude that compounds indigenous to the pineal have the potential to regulate PRL secretion from the pituitary.

D-valine medium maintains prolactin production in primary culture.

Prolactin secretion by bovine pituitary cells in L-valine-containing medium decreases approximately 96% from day 3 to day 11 of culture. We hypothesized that this decrease was caused by overgrowth of these cultures by fibroblasts. Our present objective was to maintain the synthesis and secretion of PRL by bovine pituitary cells in culture. We attempted this by growing pituitary cells in D-valine-containing medium to achieve selective suppression of fibroblast growth. Substitution of D-valine for L-valine in Earles or Swims medium resulted in undiminished PRL synthesis and release over a 30-day culture period. In contrast, comparable measures for cells maintained in medium with L-valine decreased more than 90% from day 5 to day 20 of culture and remained low thereafter. Cells cultured in medium containing D-valine retained their ability to release PRL in response to thyrotropin-releasing hormone throughout the 30-day culture period, although there was a decrease in magnitude of response with time. Similarly, estradiol increased PRL release by pituitary cells maintained in D-valine, but this stimulatory effect was no longer demonstrable by day 20 of culture. The amount of growth hormone (GH) and luteinzing hormone (LH) released into the medium decreased with time and this decrease was independent of the valine isomer contained in the medium. We conclude that substituting D-valine for L-valine in culture medium allows PRL synthesis and release to persist undiminished for at least 30 days in culture.

Effect of Arginine Vasotocin on Luteinizing Hormone and Prolactin Concentrations in Serum of Cattle

Abstract Arginine vasotocin (AVT) was given to cattle to determine its effects on concentrations of luteinizing hormone (LH) and prolactin (PRL) in serum. Doses of AVT ranging from 0.3 to 300 ü/kg BW, when given as single injection, did not alter the magnitude of frequency of normal pulsatile secretion of LH or concentrations of PRL in steers. Furthermore, AVT (0.7 mg/injection) given every 2 hr from −2 to 20 hr after 1 mg estradiol did not block the LH surge in ovariectomized cows. However, the 54% increase in concentrations of PRL which occurred 11 to 20 hr after injecting estradiol was blocked by repetitive injections of AVT. We conclude that AVT may alter PRL secretion in cattle. However, there is no indication that AVT affected LH release.