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Biomedical subjects

K J Catt

Publications and source records attributed to K J Catt.

At least 307 records · Page 17Linked to original sources
PubMed1982

Homologous regulation of gonadotropin-releasing hormone receptors in cultured pituitary cells.

Specific receptors for gonadotropin-releasing hormone (GnRH) in cultured rat pituitary cells were increased by subnanomolar concentrations of GnRH agonists and decreased by high concentrations of these peptides. The antagonist [D-Phe2, Pro3, D-Phe6]GnRH did not alter GnRH binding capacity and blocked the increase in sites induced by GnRH. These findings provide direct evidence for the homologous regulation of GnRH receptors by physiological concentrations of the hypothalamic peptide, an action that could mediate the cyclical and postcastration increases in GnRH receptors and responsiveness of the pituitary gonadotrophs.

Animals↗
PubMed1982

Characterization of angiotensin II receptors in the anterior pituitary gland.

Specific and high affinity binding sites for angiotensin II were demonstrated in the anterior pituitary gland by binding studies with [125I] iodoangiotensin II. The binding properties of the pituitary receptors were similar to those of angiotensin II receptors present in the adrenal gland. The concentration of binding sites in rat anterior pituitary (293 +/- 50 fmoles/mg protein) was less than in the adrenal gland, but was much greater than in smooth muscle. Angiotensin II receptors were identified in the anterior pituitary tissue of mature and immature animals of both sexes, and in species including rat, rabbit and dog. No binding of angiotensin II was detected in posterior pituitary homogenates, or in GH3 pituitary tumor cells. Collagenase-dispersed anterior pituitary cells also contained specific binding sites for angiotensin II, with equilibrium binding constant (Ka) of 3.6 x 10(9) M-1. The presence of specific high-affinity angiotensin II receptor in the anterior pituitary gland provides a mechanism by which angiotensin-like peptides could modulate the process of pituitary hormone secretion.

Angiotensin II↗
PubMed1982

Compartmentalization of luteinizing hormone pools: dynamics of gonadotropin releasing hormone action in superfused pituitary cells.

Cultured rat anterior pituitary cells were continuously perfused with medium 199, and exposed to short (5 min), intermediate (30 min), or long (6 h) pulses of a maximally effective concentration of gonadotropin-releasing hormone (GnRH). Assay of the effluent by radioimmunoassay and interstitial-cell bioassay revealed a biphasic response to GnRH, and indicated that 3 pools of luteinizing hormone (LH) are present in the gonadotroph. A rapidly releasable peak of bioactive LH comprising about 2% of the total cellular LH was mobilized within 1 min of GnRH addition, lasted for 3-4 min, and was independent of the duration of stimulation. The second, larger pool of bioactive LH varied from 15 to 50% of the total LH as the duration of exposure to GnRH was increased from 5 min to 6 h. A third LH pool comprising up to 50% of the total LH could be mobilized by 50 mM potassium but not by continuous GnRH treatment, due to refractoriness of the cells to prolonged stimulation by the decapeptide. In contrast, repeated pulses of GnRH evoked a series of biphasic LH peaks with profiles similar to that observed during a single response to GnRH, indicating that continuous exposure to GnRH is necessary for densensitization. Release of LH from the perfused cells was calcium-dependent, and the bio-immuno ratio of the first and second pools of LH was similar. The in vitro secretion profile of cultured rat cells is comparable with the early and late phases of LH release observed in GnRH-infused man, but occurs much more rapidly, and demonstrates heterogeneity of the LH release process at the level of the gonadotroph. The superfusion technique provides a powerful tool to further investigate the bioactivity of GnRH and its analogs for use in fertility control.

Animals↗
PubMed1982

GnRH receptors in cultured rat granulosa cells: mediation of the inhibitory and stimulatory actions of GnRH.

The regulatory actions of FSH and the GnRH agonist [D-Ala6]des-Gly10-GnRH N-ethylamide (GnRHa) upon ovarian GnRH receptors were studied in granulosa cells obtained from ovaries of hypophysectomized diethylstilbestrol-treated rats. When granulosa cells were cultured for 48 h in the presence of FSH (5-250 ng/ml) the binding of 125I-GnRHa to granulosa cell receptors was increased in a dose-dependent manner, to a maximum of 3-4 fold above the control value. Addition of FSH (100 ng) also caused a dose-dependent increase of more than 100-fold in the accumulation of cAMP and progesterone in the culture medium. In freshly prepared cells, Scatchard analysis of GnRHa binding data revealed an equilibrium constant (Ka) of 1.1 x 10(10) M-1 and GnRH receptor concentration fo 401 fmoles/mg protein. Granulosa-cell GnRH receptors decreased during culture without FSH, but were maintained in the presence of 100 ng FSH at 580 femoles/mg protein, with Ka of 0.8 x 10(10) M-1. This action of FSH on GnRH receptors was significantly reduced by 10(-8) M GnRHa. Also, GnRHa concentrations of 10(-10) and 10(-8) M caused inhibition of FSH-induced cAMP and progesterone accumulation. In cells cultured with GnRHa alone, there was a slight enhancement of GnRH receptors by GnRHa concentrations up to 10(-8) M, and a decrease below control levels with higher amounts. Also, GnRHa concentrations from 10(-8) to 10(-5) M caused a 3-4-fold increase in cAMP accumulation in the absence of FSH. These results demonstrate that FSH maintains GnRH receptors in cultured granulosa cells, and that GnRHa attenuates this effect, as well as the other actions of FSH on granulosa cell maturation. It is also evident that GnRHa itself can slightly stimulate cAMP production and partially maintain GnRH receptors, but at high concentrations causes loss of the homologous receptor sites. These findings also emphasize the ability of GnRH agonists to exert both positive and negative direct effects on rat granulosa cell function.

Animals↗
PubMed1982

Gonadotropin-releasing hormone binding and activation of enriched population of pituitary gonadotrophs.

The properties of GnRH receptors were analyzed in isolated rat pituitary cells by binding studies with the labeled GnRH agonist, [125I]iodo-[D-Ser(tBu)6]des-Gly10-GnRH-N-ethylamide. The concentration of GnRH-binding sites in pituitary cells from immature female rats was twice as high as in cells from adult females. Electron-microscopic immunocytochemistry revealed twice as many gonadotrophs in the immature rat pituitary, indicating that gonadotrophs from immature and mature female rats contain the same number of binding sites. An enriched population of gonadotrophs prepared from immature female rat pituitaries by velocity sedimentation at unit gravity contained 77% of the total radioimmunoassayable LH, and 71% of the total GnRH receptors. A second population of small, poorly granulated gonadotrophs was distributed among the cells of other fractions of the gradient, and could be detected only by immunocytochemistry. The smaller, possibly immature, gonadotrophs did not contain a measurable number of GnRH receptors. When incubated with the cells recovered from the gradient, GnRH stimulated LH release only in the mature gonadotroph-enriched fraction, and not in other regions of the gradient. These results indicate that GnRH interacts preferentially with gonadotrophs in the pituitary. The findings also suggest that functional heterogeneity exists among pituitary gonadotrophs, and that GnRH binds to and activates only the mature gonadotrophs which exhibit the highest LH-storage capacity.

Animals↗
PubMed1982

Calcium-dependent regulation of aldosterone production in isolated adrenal glomerulosa cells: effects of the ionophore A-23187.

The effect of the calcium ionophore A-23187 on the regulation of aldosterone synthesis was investigated in isolated rat adrenal glomerulosa cells. Over the concentration range of 0.1--1.0 microM, A-23187 increased aldosterone production in a dose-dependent manner, with half-maximal stimulation at approximately 0.3 microM. Stimulation of aldosterone production by A-23187 was dependent on the concentration of extracellular calcium and was not accompanied by detectable increases in cAMP production. In contrast to the stimulation of steroidogenesis elicited by A-23187 alone, the ionophore significantly inhibited the aldosterone responses to angiotensin II and potassium. In angiotensin II-stimulated cells, 0.2 microM A-23187 caused half-maximal inhibition of aldosterone production, and 2 microM A-23187 inhibited aldosterone production by about 70%. The ionophore did not affect the binding of angiotensin II to isolated glomerulosa cells and did not change the sensitivity of the cells to either angiotensin II or potassium. However, A-23187 reduced the total amount of steroid produced in response to these stimuli, most markedly when the extracellular calcium concentration was less than 0.4 mM. These results indicate that although A-23187 partially inhibits the steroidogenic responses to angiotensin II and potassium, the increases in cytosolic calcium caused by submicromolar ionophore concentrations result in stimulation of aldosterone production by adrenal zona glomerulosa cells.

Adrenal Cortex↗
PubMed1982

Binding affinity and biological activity of gonadotropin releasing hormone agonists in isolated pituitary cells.

The relationships between binding affinity and the biological potency of eight GnRH agonist analogs were evaluated in isolated rat pituitary cells. For this purpose, binding affinity and biological activity were assayed under similar physiological conditions in medium 199, pH 7.4, and binding affinity was also measured under the standard conditions in hypotonic buffer at low temperature. Under physiological conditions, receptor binding affinity was consistently lower than when measured in the hypotonic Tris buffer usually employed for GnRH receptor studies. In the low temperature binding assay at 0 C, which provided a measure of the affinity constant without degradation, a difference of 20- to 30-fold was observed between native GnRH and its most potent analog, [D-Ser(t-Bu)6]des-Gly10-GnRH N-ethylamide. Modifications of the amino acid residues at both positions 6 and 10 of the decapeptide increased the binding affinity of GnRH analogs. When the receptor binding assay was performed at 37 C, the range of the apparent affinity constants was extended up to 60-fold. The affinity constants derived at 37 C were closely correlated with the biological potencies of the individual analogs measured in the same cell system. The effect of temperature on binding affinity was not significantly influenced by peptide metabolism, which was minor in the absence of horse serum from the incubation medium. At the pituitary level, the biological potency of the GnRH agonist analogs is predominantly determined by their higher receptor affinity, and reduced degradation is a less important aspect of the high biological activity of the superagonist analogs.

Animals↗
PubMed1982

Gonadotropin binding and Leydig cell activation in the rat testis in vivo.

Testicular LH receptor occupancy and steroidogenic responses were measured in adult male rats after intracardiac injections of [125I]iodo-hCG (0.5--5 x 10(6) cpm) mixed with known amounts of nonradioactive hCG to yield doses ranging from 10 ng to 300 micrograms. Uptake of the hormone by the testis was measured in the whole tissue or the 20,000 x g homogenate, with correction for nonspecific binding in animals injected with a 100-fold excess of unlabeled hCG. The steroidogenic response to hCG was followed by measurements of serum and testicular testosterone. Maximum specific uptake of [125I]iodo-hCG by the testes was observed 4--6 h after hormone injection. Of the specific counts, 80% were recovered in the 20,000 x g pellet of the tissue homogenate. The testicular contents of hCG-binding sites were similar when measured by in vivo occupancy of the receptors and by the in vitro receptor assay, indicating the physiological validity of the receptor measurements in tissue homogenates. Serum and testicular testosterone levels reached a maximum at 1 h, independent of the hCG dose used. When receptor occupancy in vitro after injection of hCG was compared with stimulation of steroidogenesis, a significant (P less than 0.05) 3-fold elevation of serum testosterone was seen when only 0.05% of the receptors were occupied. The maximal testosterone response was reached with 0.8% receptor occupancy. It is concluded that the same number of testicular LH receptors can be occupied by the circulating hormone in vivo and in tissue homogenates in vitro. The spare receptor concept also applied to the in vivo situation, since stimulation of steroidogenesis in the intact animal requires occupancy of only a few receptors per Leydig cell. This may be a general feature of hormonal activation of endocrine target cells in vivo.

Animals↗
PubMed1982

Angiotensin II receptors and prolactin release in pituitary lactotrophs.

Logical properties of angiotensin II receptors in the rat adenohypophysis were analyzed in cultured rat pituitary cells incubated with angiotensin II and known stimuli of pituitary hormone secretion. PRL release during incubation for 3 h with 3 nM angiotensin II was consistently increased by 68 +/- 5%, comparable with that elicited by TRH (63.1 +/- 4%). The ED50 of 0.5 nM for PRL release by angiotensin II was significantly lower than that of TRH (2.9 nM) in the same cell cultures. The antagonist analog [Sar1,Ala8]angiotensin II prevented the angiotensin-induced rise in PRL production but not that evoked by TRH, whereas dopamine and SRIF inhibited basal, angiotensin, and TRH-stimulated PRL release. Angiotensin II also caused a small increase in ACTH release but had no effect on the release of LH, TSH, and GH. Angiotensin II binding and PRL release were measured in partially purified lactotrophs prepared by elutriation, by which the initial cell suspension was separated into seven fractions. Most of the lactotrophs were present in the two fractions eluted at flow rates of 15.7 and 19.8 ml/min, as indicated by their immunoreactive PRL content. The 2.5- to 3.2-fold enrichment of lactotrophs was accompanied by a 2- to 3.5-fold increase in angiotensin II receptor concentration, with no change in binding affinity (Ka = 3.5 x 10(9) M-1). In the same fractions, angiotensin II-induced PRL release was similarly increased by 1.6- to 3.5-fold above basal, compared with values of less than 1 in the initial cell suspension and other fractions. The preferential location of angiotensin II receptors in the lactotroph-containing fractions and the close correlation between angiotensin II binding sites and stimulation of PRL release indicate the functional importance of the pituitary angiotensin II receptor sites. These findings also suggest that angiotensin II could contribute to the physiological regulation of PRL secretion.

Angiotensin II↗
PubMed1982

Characterization of somatostatin receptors in the rat adrenal glomerulosa zone.

Specific receptors for somatostatin have been identified and characterized in the rat adrenal glomerulosa zone in vivo and in vitro by binding studies with [125I]iodo-Tyr-somatostatin. In adult rats, the injection of [125I]iodo-Tyr-somatostatin was followed by rapid uptake of the labeled peptide in several tissues. The highest uptake was in the adrenal capsule, with a tissue to blood ratio of 4.5, followed by kidney, anterior pituitary, and liver with tissue to blood ratios of 2.0, 1.8, and 1.4, respectively. In vitro binding studies with adrenal capsular particles were performed at 16 C in the presence of bacitracin and thimerosal to reduce tracer degradation. Under these conditions, binding of [125I]iodo-Tyr-somatostatin to capsular membrane-rich fractions reached a steady state within 30-40 min and remained at a plateau for 120 min. Dissociation of bound somatostatin from its adrenal receptors was also rapid, with an initial half-time of 5 min. Equilibrium binding of somatostatin to adrenal capsular particles was saturable and of high affinity, with an association constant (Ka) of 1.5 x 10(10) M-1. The receptor-binding activities of several somatostatin analogs were consistent with their potencies as inhibitors of angiotensin II-stimulated aldosterone production in adrenal capsular cells and with their known biological activities upon GH release. These findings demonstrate that high affinity receptors with structural and biological specificities for somatostatin are present in the adrenal glomerulosa zone. Such receptors serve as the regulatory sites through which somatostatin inhibits the action of angiotensin II upon aldosterone production in the adrenal glomerulosa cell.

Adrenal Glands↗