Platelet-derived growth factor modulates epidermal growth factor receptors by a mechanism distinct from that of phorbol esters.

Platelet-derived growth factor (PDGF) and phorbol 12-myristate 13-acetate (PMA) decrease high affinity binding of 125I-labeled epidermal growth factor (EGF) and potentiate mitogenesis in BALB/c 3T3 cells, and both have been shown to induce the phosphorylation of the EGF receptor at threonine residues. These similarities suggest that the actions of PDGF on EGF binding may be mediated by protein kinase C, the cellular effector of PMA. We show that in density-arrested BALB/c 3T3 cells PDGF and PMA induce a rapid, transient, cycloheximide-independent loss of EGF binding activity. As has been previously shown for PDGF, the ability of PMA to reduce EGF binding was enhanced by cholera toxin, a potent activator of adenylate cyclase. In contrast to PMA, however, PDGF induced a further reduction in EGF binding that was strictly dependent upon continued protein synthesis. Furthermore, PDGF effectively reduced EGF binding in cells refractory to PMA. Cells desensitized to PMA, presumably due to the loss of protein kinase C activity, also remained mitogenically responsive to PDGF. These data suggest that the mechanism by which PDGF modulates EGF binding differs from that of PMA and thus, at least in part, is independent of protein kinase C.     

Evidence that opiatergic and alpha-adrenergic mechanisms stimulate rat growth hormone release via growth hormone-releasing factor (GRF)

An antiserum raised against human pancreatic growth hormone-releasing factor-(1-40) (hpGRF-40) was found to recognize the rat hypothalamic growth hormone-releasing factor (rhGRF): This antiserum, when given iv to adult male rats, completely abolished GH release induced by a synthetic rhGRF (0.2, 1 microgram/kg BW), and inhibited the physiological GH secretion over 2 days. After passive immunization with anti-hpGRF-40 serum, iv injection of either FK 33-824 (100 micrograms/kg BW), an enkephalin analogue, or clonidine (125 micrograms/kg BW), an alpha-adrenergic agent, failed to stimulate GH release in the unanesthetized rat. These results indicate that the GH-releasing actions of opiatergic and alpha-adrenergic mechanisms are mediated through stimulation of endogenous rhGRF release in the rat.     

A possible role of prostacyclin to stimulate prolactin and growth hormone release by hypothalamic and pituitary actions, respectively

Prostacyclin (PGI2) (1-5 micrograms in 3 microliters 0.05 M Tris/HCl buffer, pH 7.5) and its stable metabolite, 6-oxo-PGF1 alpha, were microinjected into the third ventricle of ovariectomized rats, and plasma FSH, GH, PRL, and TSH levels were measured by RIA. Control animals received 3 microliters buffer. Injection of 5 micrograms PGI2 dramatically elevated plasma PRL values (4- to 5-fold) at 5 and 15 min, whereas the same dose of 6-oxo-PGF1 alpha produced a significant but smaller (2-fold) stimulatory effect. A delayed increase (1.5-fold) in plasma GH occurred after intraventricular PGI2 at 30 and 60 min. 6-Oxo-PGF1 alpha failed to alter GH levels. There were no alterations in plasma FSH and TSH after intraventricular injection of PGI2. Dispersed, overnight cultured cells from anterior pituitaries of ovariectomized rats were tested with 10(-4)-10(-7) M PGI2 and its metabolite. After 15 min of incubation, 3 X 10(-5) PGI2 produced a highly significant elevation in GH release (P less than 0.001), whereas there was no alteration in PRL levels. Only pharmacological doses of 6-oxo-PGF1 alpha (10(-4) M) stimulated GH release. There was no alteration in PRL release by the cultured cells even in the presence of 10(-4) PGI2. These results suggest that PGI2 stimulates PRL release by a hypothalamic action either to increase the release of PRL-releasing factor, or to decrease release of PRL-inhibiting factor, or by both mechanisms. The delayed stimulatory effect of PGI2 on the release of GH may be exerted via an effect on the anterior lobe itself, since PGI2 was effective in stimulating GH release by the incubated pituitary cells.     

Correlation between hormone dependency and the regulation of epidermal growth factor receptor by tumor promoters in human mammary carcinoma cells.

The effects of the tumor promoter phorbol 12-tetradecanoate 13-acetate (TPA) on the epidermal growth factor (EGF) receptor levels were investigated in hormone-dependent (MCF-7, T-47-D, and ZR-75-1) and hormone-independent (MDA-MB-231, HBL-100, and BT-20) human mammary carcinoma cell lines. In the absence of TPA, hormone-independent cell lines contained high concentrations of low-affinity EGF receptors (apparent Kd = 8 X 10(-10) M), whereas hormone-dependent cell lines exhibited low concentrations of high-affinity receptors (apparent Kd = 1 X 10(-10) M). TPA causes a change of the receptor from a high- to the low-affinity state in hormone-dependent cell lines (MCF-7, T-47-D, and ZR-75-1), as well as in the hormone-independent HBL-100, whereas the affinity remained unchanged in MDA-MB-231 and BT-20 cells. In addition, progesterone receptor levels are decreased after TPA treatment in the hormone-dependent cell lines MCF-7, T-47-D, and ZR-75-1, whereas the estrogen receptor levels remained unchanged. Tumor promoters such as TPA or teleocidin inhibited the proliferation of these cell lines at concentrations above 10 microM with the exception of the T-47-D cells. The most sensitive cell line towards growth inhibition by tumor promoter was the hormone-dependent MCF-7 cell line. Evaluation of different TPA analogs indicated a positive correlation between the growth-inhibitory effects and their ability to stimulate the subcellular redistribution of protein kinase C activity in MCF-7 cells. These data suggest a protein kinase C-mediated down-regulation of the progesterone receptor concentration and of the EGF receptor affinity, which is supposed to mediate the mitogenic response. Furthermore, these results support the hypothesis that the tumor-derived growth factors induced by estradiol act via the EGF receptor in hormone-dependent mammary carcinoma cells.     

The 20-kilodalton (kDa) human growth hormone (hGH) differs from the 22-kDa hGH in the effect on the human prolactin receptor.

Previously we have demonstrated that 20-kDa human GH (20K-hGH) is a full agonist for hGH receptor (hGHR) even though its complex formation with hGHR and hGH-binding protein differs from that of 22-kDa human GH (22K-hGH). In this study, we focused on the effect of 20K-hGH on human PRL receptor (hPRLR). To elucidate the effects of 20K-hGH on hPRLR and compare them with those of 22K-hGH, we prepared two cells stably expressing full-length hPRLR, Ba/F3-hPRLR and CHO-hPRLR. In the proliferation of Ba/F3-hPRLR cells, which can grow in a dose-response to lactogenic hormones, both 20K- and 22K-hGH exhibited bell-shaped curves in the absence of exogenous zinc ion (Zn2+); however, the curve of 20K-hGH was shifted to a 10-fold higher concentration than that of 22K-hGH in view of EC50 value (the EC50 of 20K- and 22K-hGH were 15 nM and 1.5 nM, respectively). Addition of Zn2+ up to 25 microM increased the activities of both 20K- and 22K-hGH; however, the enhancement by Zn2+ was greater in 20K-hGH than in 22K-hGH, thereby the activities of both hGH isoforms reached the same level at 25 microM Zn2+. Nevertheless, in the presence of 0.25-1 microM free Zn2+, which is equal in human serum, the activity of 20K-hGH was still lower than that of 22K-hGH. The modest effect of 20K-hGH on activating hPRLR in the absence of Zn2+ was confirmed in the rat serine protease inhibitor 2.1 (Spi2.1) gene promoter activation and JAK2/Stat5 tyrosine phosphorylation in CHO-hPRLR. In addition, in human breast cancer cell T-47D, 20K-hGH was proved to stimulate Stat5 tyrosine phosphorylation to much lower degree than 22K-hGH via not hGHR but hPRLR. Taken together, our data suggest that 20K-hGH may be a weaker agonist for hPRLR than 22K-hGH in the human body; therefore 20K-hGH may alleviate the hPRLR-mediated side-effects such as breast cancer when administered to human body.     

Evidence that phorbol diester-sensitive protein kinase-C(s) may not be directly involved in secretagogue-stimulated prolactin release and arachidonate liberation

This report presents findings pertaining to the role of protein kinase-Cs in the release of PRL and liberation of arachidonate from PRL-secreting cells. In our experiments, protein kinase-C activators increased PRL release and arachidonate liberation from anterior pituitary cells and from the PRL-secreting cell line MMQ. In cells depleted of pituitary protein kinase-Cs by chronic exposure to protein kinase-C activators, such as phorbol dibutyrate or 4 beta-phorbol 12 beta-myristate 13 alpha-acetate, TRH, angiotensin-II, and neurotensin each increased PRL release and [3H]arachidonate liberation in a normal manner. In addition, the PRL-releasing activities of protein kinase-C activators and those of TRH appeared to be synergistic, an unexpected effect if these substances were functioning through the same intracellular pathways. It, therefore, appears that phorbol diester-sensitive protein kinase-Cs may not be involved in the increased secretion of PRL or liberation of arachidonate that is caused by TRH, angiotensin-II, or neurotensin.     

Effects of epidermal growth factor, phorbol ester, and retinoic acid on hormone synthesis and morphology in porcine thyroid follicles cultured in collagen gel.

Epidermal growth factor (EGF), phorbol esters (PEs), and retinoic acid (RA) inhibit differentiated functions of thyrocytes. In the present study the inhibitory effects of these growth-promoting factors on hormone synthesis were studied in thyroid follicles cultured in type-I collagen gel, and morphologic alteration by these factors was examined by light and electron microscopy (EM). Porcine open thyroid follicles obtained by treatment with 0.1% collagenase were embedded in collagen gel and cultured in Ham's F12 medium supplemented with 6H (insulin, hydrocortisone, somatostatin, transferrin, glycyl-his-lys, and thyrotropin) + 0.5% fetal bovine serum (FBS). After 1 week these open follicles developed to closed follicles, and the medium was changed to one containing 6H + 0.5% FBS + 0.1 microM sodium iodide (NaI). Some media were supplemented with either EGF, phorbol 12-myristate 13-acetate (PMA), or all-trans RA. The closed follicles retained ability for hormone synthesis for 2 weeks after the medium change in the presence of 6H + FBS + NaI. The amounts of T4 and T3 secreted into the culture medium from day 9 to day 12 after the medium change were 60% and 45% of those from day 0 to day 4, respectively. EGF reduced production of T4 and T3 by 61% and 69%, respectively; PMA, by 87% and 99%; and RA, by 55% and 44%. In the medium supplemented with 6H + 0.5% FBS, the follicles exhibited intact polarity. Apical surfaces with microvilli were oriented to the follicular lumen and tight junctions were on the apical side of cell-to-cell contacts. Desmosomes were found on both the apical and basal halves of the cell contacts.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of epidermal growth factor on insulin-like growth factor-I (IGF-I) and IGF-binding protein synthesis by adult rat hepatocytes.

Growth hormone has been established as a primary regulator of IGF-I gene expression in adults, not only in liver but also in many extrahepatic tissues. We considered the possibility that IGF-I production by adult rat liver could also be stimulated by epidermal growth factor (EGF), a peptide known to be involved in liver regeneration. Chromatographic analysis performed after acid treatment of conditioned media revealed the presence of both immunoreactive (IR) IGF-I and IGF binding protein (IGFBP). Both IR IGF-I and IGFBP were present in the conditioned medium of adult rat hepatocytes in basal conditions. The stimulation of IGF-I and IGFBP secretion by EGF appears to be dose-dependent with a significant increment already evident at 5 nM. That EGF stimulates secretion is supported by the finding that IGF-I and IGFBP-1 mRNA levels are increased after EGF supplementation. We conclude that adult rat hepatocytes spontaneously produce IGF-I and IGFBP, and that EGF is able to increase their synthesis and secretion. This non-growth hormone-dependent regulation of IGF-I and IGFBP-1 production by adult rat hepatocytes in culture indicates an important autocrine/paracrine role for IGF-I, particularly during liver regeneration after extensive organ mass loss.     

Vesicular release of prolactin from preformed prolactin granules is stimulated by soluble factor(s) from the anterior pituitary of lactating rats

This study demonstrates that conditioned media (CM) from the anterior pituitary gland (AP) of lactating rats contains soluble factors that promote in vitro prolactin (PRL) release from the pituitary glands of male rats. CM-induced PRL release was confirmed by polyacrylamide gel electrophoresis, ELISA and bioassay. In cultured AP cells challenged with CM, increased intracellular staining with the dye FM1-43 was observed, suggesting vesicular PRL release and subsequent endocytosis. The percentage and hormone content of PRL-containing cells but not of growth hormone-containing cells increased in cultured male AP cells when exposed to CM. When the release of PRL, prelabeled with [3H] leucine for 30 min to 24 h was examined, no stimulatory effect of CM was observed, suggesting that released PRL originates from hormone synthesized more than 24 h earlier. Accordingly, the PRL content of mature granules from male pituitary tissues decreased after CM treatment. These findings were confirmed by electron microscopy immunogold PRL labeling. Treatment with inhibitors of protein synthesis or vesicle trafficking between the endoplasmic reticulum and the Golgi complex did not prevent the stimulatory effect of CM on PRL release. However, blockage of traffic to the plasma membrane completely abolished the effect of CM. These results suggest that CM from the AP of lactators contains soluble factor(s) capable of inducing rapid vesicular release of PRL in the male AP, which originates from preformed, mature granules by mechanisms independent of protein synthesis.     

Simultaneous detection of epidermal growth factor receptor (EGF-R), epidermal growth factor (EGF) and ras p21 in cholangiocarcinoma by an immunocytochemical method

Expression of the epidermal growth factor (EGF), EGF receptor (EGF-R) and ras oncogene product p21 was simultaneously examined in 37 cases with intrahepatic cholangiocarcinoma (CC) by means of an immunocytochemical method. While normal livers were all negative for any of the antigens at the concentration of the antibodies used, EGF-R was positive in 12 (32.4%) CCs, EGF in 22 (59.5%), and ras p21 in 33 (89.2%). The positive incidence of the three antigens was not different among the histologic subtypes of the tumor. However, the number of EGF-R- and ras p21-positive tumor cells decreased with progressing histologic tumor grade, but the expression of EGF was not associated with the tumor grade. Expression of the three antigens was not related to the degree of metastatic spread of the tumor. Simultaneous expression of the three antigens was seen only in 4 CCs, and that of EGF-R and EGF in 4, EGF-R and ras p21 in 12, and EGF and ras p21 in 20. These data suggest that the expression of EGF, EGF-R and ras p21 on CC cells is not related to the tumor aggressiveness, and the activation of each respective gene is independent. Furthermore, the data also indicate that an autocrine model for tumor growth, as suggested by a combination of EGF and EGF-R, may be applicable only to very limited cases of CCs.     

The effect of epidermal growth factor (EGF) with estradiol and progesterone on prolactin secretion from cultured human decidual tissues of early pregnancy

Prolactin is now accepted as a normal product of the decidual cells of the human endometrium. We investigated the effect of epidermal growth factor (EGF) with estradiol and progesterone on prolactin secretion by the decidual tissues from the early pregnant endometrium. The decidual tissues were separated from villi, minced and cultured in collagen gel matrix with serum-free medium. Immunological staining of the cultured decidual tissues showed prolactin localization and EGF receptors on the stromal cells. Cultured media were collected every 2 days. The culture for the first 2 days was incubated with the serum-free medium alone (= preculture), and the following test culture was supplemented with/without additives. The prolactin content in cultured media was quantified by EIA. The results of the effect of steroid(s) and EGF were represented as a comparison of prolactin contents in the preculture and the test culture. An increase in prolactin secretion was found after the tissues were treated with a combination of 10(-8)M estradiol and 10(-6)M progesterone or 10(-6)M progesterone alone. After 8 days, the prolactin secretion rate increased about 3-fold over the precultured value. Estradiol alone kept the prolactin secretion at the precultured value. Prolactin secretion gradually decreased in the non-additive culture. These results indicate that progesterone was essential in the secretion of prolactin. Simultaneously, similar decidual tissues were incubated with a combination of EGF and steroid(s). The secretion of prolactin in the group treated with progesterone alone decreased dose-dependently responding to added EGF on the 8th day of culture. In the presence of estradiol and progesterone, the secretion rate decreased to the values similar to the progesterone alone group with the addition of 0.1, 1 ng/ml EGF, and the decrease in prolactin secretion was less with the addition of 10 ng/ml EGF. Mixed cultures of the decidual tissues and villi showed that the prolactin secretion rate increased in all groups treated with/without estradiol and/or progesterone. These results imply that progesterone derived from villi might control decidual prolactin secretion. The effect of high concentration EGF (50 ng/ml) on the prolactin secretion appeared similar to the isolated decidual tissues. These results suggest that decidual prolactin secretion is regulated by the combined effects of steroids and EGF.     

Regulation of surface expression of high-affinity receptors for epidermal growth factor (EGF) in hepatocytes by hormones,differentiating agents,and phorbol ester

Freshly isolated adult rat hepatocytes exhibit a nonhomogeneous population of epidermal growth factor (EGF) receptors with about 10,000 high-affinity binding sites (Kd 20 pM) and about 200,000 low-affinity sites (Kd 600 pM) per cell. With culturing as primary monolayers under conditions where the cells show a marked increase in the sensitivity to the growth-stimulatory effect of EGF, a gradual reduction in the number of EGF receptors and an almost complete loss of high-affinity EGF receptors is seen. Insulin, which promotes growth of hepatocytes in concert with EGF, enhances the down-regulation of these high-affinity receptors. The differentiating (and growth-inhibitory) agent n-butyrate counteracts this down-regulation and preserves the high-affinity receptors. This effect of butyrate is synergistic with the glucocorticoid agent dexamethasone. Another differentiating agent, dimethylsulfoxide (DMSO), also counteracts the down-regulation of high-affinity EGF receptors. Moreover, the tumor promoter, tetradecanoylphorbol acetate (TPA), down-regulates the EGF receptor. This effect is particularly evident when studying the high-affinity receptors up-regulated by prior treatment with butyrate plus dexamethasone. Taken together these results provide strong support for the notion that an inverse relationship exists between expression of high-affinity EGF binding and responsiveness to growth activation by EGF.     

Heparin-dependent binding and autophosphorylation of epidermal growth factor (EGF) receptor by heparin-binding EGF-like growth factor but not by EGF.

Heparin-binding EGF-like growth factor (HB-EGF) is a recently identified member of the EGF family of growth factors and a potent mitogen for smooth muscle cells and fibroblasts. Chinese hamster ovary (CHO) cells genetically engineered to express the human EGF receptor bind with high affinity both EGF and HB-EGF. CHO mutant cells lacking heparan sulfate proteoglycans (HSPG) bind EGF equally well to wild-type cells and EGF binding is not affected by exogenous heparin. However, HSPG-deficient EGF receptor-expressing cells do not bind significant levels of HB-EGF unless heparin is present in the binding medium. Moreover, binding of radiolabeled EGF to HSPG-deficient EGF receptor-expressing cells is efficiently displaced by nonlabeled HB-EGF only in the presence of heparin. Signal transduction by the EGF receptor tyrosine kinase as evidenced by receptor autophosphorylation is induced by HB-EGF only in the presence of heparin, in contrast to EGF-induced receptor autophosphorylation, which is independent of heparin. These results directly demonstrate that HB-EGF but not EGF requires heparin or cell surface HSPG for binding and activation of the EGF receptor and that HB-EGF receptor interactions can be tightly regulated by the available local concentration of heparin-like molecules.     

Growth hormone and prolactin stimulate androgen receptor, insulin-like growth factor-I (IGF-I) and IGF-I receptor levels in the prostate of immature rats.

In this study we investigated the involvement of several different pituitary hormones on rat prostate development. 22-day-old Wistar rats, hypophysectomized (hypox) at 19 days of age were supplemented with highly purified human prolactin (hPRL), human luteinizing hormone (hLH), porcine follicle-stimulating hormone (pFSH), and bovine growth hormone (bGH) or with saline. Quantitative analysis of RNAs shows that treatment with either PRL or GH increases significantly steady-state mRNAs levels of the following genes in the prostate: androgen receptor (AR) (respectively 3.5- and 4.8-fold above hypox controls), IGF-I (5- and 2.7-fold), and IGF-I receptor (2.9- and 2.3-fold). LH and FSH, by contrast, have negative effects on these parameters. To test whether the enhancing effect of PRL and GH on AR-mRNA abundance was followed by increased content in the protein itself, binding assays were performed with the androgen agonist [3H]R1881 (131 and 153 fmol/mg protein while hypox controls contained 110 fmol/mg protein). In addition to the well-documented presence of prolactin receptors in prostatic tissues, we have further demonstrated, by means of nuclease S1 protection assays plus dot- and Northern-blot analyses, that a GH receptor mRNA is produced in the immature rat prostate. Moreover, we observed not only strong lactogenic but also purely somatogenic binding to be occurring in the immature prostates. Finally, we have studied IGF-I mRNA content in separated epithelial/stromal cell fractions and have concluded that IGF-I expression is principally located in the prostatic stroma. Taken together, these results suggest that PRL and GH are involved in regulating AR synthesis, at least partially by direct action on the organ. In this context IGF-I appears as a paracrine factor playing a role in epithelium/stroma interactions during prostatic development.     

Regulation of epidermal growth factor receptors by TSH, and effects of EGF, TSH and phorbol ester on DNA synthesis in cultured porcine thyroid cells

Using porcine thyroid cells of primary monolayer culture, this study was conducted to clarify the nature and characteristics of epidermal growth factor (EGF) receptors on porcine thyroid cells and also to investigate the effects of EGF, TSH and phorbol ester on DNA synthesis. Receptors for EGF on porcine thyroid cells exist in two forms which differ in both affinity and capacity. Scatchard analysis of saturation binding assay performed at 4 degrees C for 6h indicates that there is a high affinity class of receptors with low capacity (K1 = 4.70 +/- 0.90 X 10(-9)M and 8,600 +/- 1,200 sites/cell) and low affinity receptors with high capacity (K2 = 2.24 +/- 1.17 X 10(-7)M and 65,500 +/- 18,000 sites/cell) on the cells cultured for 4 days in the absence of TSH. When thyroid cells were cultured in the presence of various concentrations of TSH (0 approximately 50 mU/ml) and for various times (0 approximately 96 h) with TSH (10 mU/ml), specific EGF binding to the cells increased dose- and time-dependently. On TSH (10 mU/ml)-treated cells for 4 days, two kinds of EGF receptors, i.e. high affinity and low capacity (K1 = 5.39 +/- 1.75 X 10(-9) M and 17,200 +/- 2,500 sites/cell) and low affinity and high capacity (K2 = 1.70 +/- 1.40 X 10(-7)M and 76,300 +/- 17,900 sites/cells), were resolved. The results indicate that TSH can modulate EGF receptors by increasing the number of high affinity sites on porcine thyroid cells. Next, using [Me-3H] thymidine incorporation into TCA precipitable materials for 48h, we studied the biological effects of EGF, TSH and phorbol myristate acetate (one of the potent phorbol esters) on DNA synthesis. Both EGF and PMA can promote [Me-3H] thymidine incorporation. Maximal responses were obtained with EGF ranging from 10(-9) to 10(-7)M and with PMA from 10(-9) to 10(-7)M. In contrast, TSH inhibits [Me-3H] thymidine incorporation dose-dependently. Almost the same results were obtained by these agents on TSH (10 mU/ml)-treated cells for 4 days, but EGF stimulated cell growth at a lower concentration of 10(-11)M, which was possibly related to an increase of high affinity receptor numbers. The growth promoting effect of EGF and PMA in combination was additive, and TSH suppressed the cell growth in the concomitant presence of EGF and PMA.(ABSTRACT TRUNCATED AT 400 WORDS)     

Comparative stimulation of growth hormone secretion in anaesthetized chickens by human pancreatic growth hormone-releasing factor (hpGRF) and thyrotrophin-releasing hormone (TRH)

Plasma concentrations of growth hormone (GH) were elevated in anaesthetized male domestic fowl following the intravenous administration of either synthetic human pancreatic GH-releasing factor 1-44 (NH2) (hpGRF) or synthetic thyrotrophin-releasing hormone (TRH). In 6-week-old chicks the plasma GH level was elevated between 5 and 10 min after the injection of hpGRF at doses between 1 and 80 micrograms/kg. The magnitude of the response increased with doses of hpGRF between 1 and 10 micrograms/kg but declined with higher doses. The GH concentration rapidly declined between 10 and 20 min and between 20 and 40 min after injection. The administration of TRH had similar effects on GH secretion, although the responses were greater than with comparable doses of hpGRF, and the most effective dose (1-1.4 micrograms/kg) was less than with hpGRF. In anaesthetized adult cockerels GH secretion was also increased by the administration of hpGRF (1-20 micrograms/kg) or TRH (0.1-80 micrograms/kg) and in both cases the dose-response relationship was biphasic. The maximal response to TRH in adult birds was again greater than that produced by hpGRF although the response was less than that elicited in immature birds and required a higher dose (20 micrograms/kg) of TRH. The optimal dose of hpGRF and the magnitude of the GH response induced in adult birds was comparable with that in immature chicks. These results demonstrate provocative effects of TRH and hpGRF on GH secretion in the domestic fowl. The sensitivity of the GH response to TRH suggests that it may have a physiological role in the hypothalamic control of GH secretion.