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The major anionic mitogenic polypeptide for endothelial cells, acidic fibroblast growth factor (FGF), has been purified to homogeneity from bovine brain and its complete primary structure established by gas-phase sequence analysis. The 140 amino acid (Mr 16,000) protein has been previously shown to be a potent growth factor for many diverse cell types of mesodermal origin, in vitro, and an angiogenic agent, in vivo. The amino acid sequence of bovine brain acidic FGF has a 53% absolute homology with that of bovine pituitary basic FGF suggesting that these endothelial cell mitogens are derived from a single ancestral gene.

Summary While showing consistent and easily reproducible results as observed in vitro or in acute in vivo systems, several N α -acylated-des-[Ala 1 -Gly 2 ]-dihydrosomatostatin analogs exhibit variable protracted inhibition of the growth hormone release induced in rats by pentobarbital. These results may reflect variable central nervous system responses to pentobarbital as well as to the somatostatin analogs.

A peptide with somatostatin-like immuno- and bioactivity has been isolated from an aqueous extract of 96 800 rat hypothalami by means of immunoaffinity chromatography, gel filtration and reverse-phase high-performance liquid chromatography (HPLC). Chemical characterization by amino acid analysis, tryptic peptide mapping and retention behavior in two HPLC systems showed that the peptide was indistinguishable from somatostatin-28 previously characterized from several species. This evidence suggests that rat hypothalamic somatostatin-28 is identical in structure to porcine and ovine somatostatin-28.

We have isolated from extracts of ovine hypothalami two molecules characterized as somatostatin-28 and somatostatin-4–28 (referred to as somatostatin-25). They were reproduced by solid phase synthesis. In equimolar ratio and depending upon the experimental conditions, synthetic somatostatin-28 and somatostatin-25 are 3–14 times more potent than somatostatin-14 to inhibit the basal in vitro secretion of growth hormone or as stimulated by prostaglandin (PGE 2 ). In early studies in vivo, somatostatin-28 and somatostatin-25 are also more potent than somatostatin-14 in inhibiting the secretion of growth hormone acutely stimulated in the rat by injection of morphine; somatostatin-28 is also longer-acting than somatostatin-14. These results suggest that somatostatin-14, as originally isolated, is a biologically active fragment of a larger molecule of greater specific activity; it should be considered as another form of somatostatin with high biological activity present in some tissues and likely secreted by the tissues along with somatostatin-14 and possibly other somatostatin-peptides of diverse sizes.

Human hypothalamic growth hormone-releasing factor (GRF) was purified by gel filtration and reverse-phase HPLC. Bioassay and two radioimmunoassays of different specificity revealed the presence of two major forms of GRF-activity which coelute with human pancreas GRFs, hpGRF-44-NH2 and hpGRF-40 previously characterized in pancreas tumors. The bioactive material coeluting with hpGRF-44-NH2 is recognized by two antibodies which are directed against the amidated COOH-terminal sequence and the central portion of the GRF-44 peptide. The bioactive GRF which coelutes with hpGRF-40 reacts only with the antibody directed against the central portion of hpGRF. These data strongly suggest that the human hypothalamus contains the same major forms of GRF that were identified in pancreas tumors responsible for acromegaly in the absence of a pituitary tumor.

Two mitogenic peptides in bovine liver extract were purified to apparent homogeneity by monitoring the purification steps with two in vitro bioassays; one based on stimulation of adult bovine aortic arch endothelial cell proliferation and the other incorporation of [ 3 H]thymidine to mouse fibroblast 3T3 cells. The purification procedure involved cation-exchange chromatography followed by affinity chromatography on heparin-Sepharose and two steps of reversed-phase HPLC. The purified material showed the same biological activity as pituitary basic fibroblast growth factor (FGF). Amino acid analyses of the purified mitogen yielded a similar, but not identical composition to that of bovine pituitary basic FGF(1–146) reported previously. Gas-phase microsequencing identified two sequences in equal amounts in the purified preparation. Furthermore, the sequencing results are in accord with the theoretical data obtained when two truncated forms of basic FGF, corresponding to FGF(12–16) and (16–146), are being sequenced simultaneously. Basic FGF(12–146) is a novel truncated form of basic FGF which has not been isolated before although the (16–146) fragment has been found previously in kidney, corpus luteum, and adrenal. SDS-PAGE analysis could not separate the two forms and showed that both migrated as a protein of about 15 100 daltons, which is slightly smaller than intact basic FGF(1–146) (16 200 daltons). These results, taken together, indicate that at least some of the mitogenic activity in liver may be derived from basic FGF-related polypeptides.

The synthetic replicate of a 44 amino acid peptide isolated from a human pancreatic tumor which had caused acromegaly possesses high specific activity to release growth hormone (GH) in anesthetized male rats. The GH secretion induced by this peptide is dose-dependent from 50 ng to 1 μg, with plasma GH concentrations increasing more than 10-fold within 5 min of iv administration at the higher doses. Two enzymatic degradation products of the 44 residue peptide were also isolated and consist of the first 37 and 40 amino acids. All three peptides appear to possess similar potency, on a molar basis, invivo, contrary to invitro results. The specificity of these peptides on GH release was shown by their failure to alter plasma concentrations of prolactin (PRL), thyroid-stimulating hormone (TSH), luteinizing hormone (LH), follicle-stimulating hormone (FSH) and corticosterone. Based on these invivo results, the three peptides with serve as powerful tools with which to investigate the mechanisms of GH secretion.

The brain levels of β-endorphin, α-endorphin and enkephalin were measured by radioimmunoassay after different methods of sacrifice. Microwave irradiation proved not to be better than decapitation followed by boiling of the intact tissue, the latter procedure giving values of β-endorphin 10 fold higher than decapitation alone. Concurrently when decapitation was followed by boiling, α-endorphin was no longer detected. Evaluation in brain tissue of several extraction media--phosphate buffered saline, 5% TCA, HCl methanol, and 1N HOAc--showed the last to be the most satisfactory for both β-endorphin and enkephalin. Since β-endorphin was found to be readily hydrolized by brain homogenates with consequent appearance of α-endorphin, these results indicate that disruption of tissue modifies the content of opioid peptides in brain.

A 44 amino acid peptide with high intrinsic growth hormone releasing activity was isolated from 500 bovine hypothalami by means of acid extraction, immunoaffinity chromatography, gel filtration, and two steps of reverse phase HPLC. The growth hormone releasing factor was structurally characterized by gas phase sequence analysis. Reverse phase liquid chromatography of the native peptide and synthetic replicates showed that the molecule possesses an amide rather than a free acid at its carboxyl terminus. The structure of the peptide was established as: Tyr Ala-Asp-Ala-Ile-Phe-Thr-Asn-Ser-Tyr-Arg-Lys-Val-Leu-Gly-Gln-Leu-Ser-Ala -Arg-Lys-Leu-Leu-Gln-Asp-Ile-Met-Asn-Arg-Gln-Gln-Gly-Glu-Arg-Asn-Gln -Gly-Ala-Lys-Val-Arg-Leu-NH2 using approximately 2 nmol of material.

The primary structures of three polypeptides, possessing high intrinsic growth hormone-releasing activity and derived from a human pancreatic carcinoma which had caused acromegaly, were established by sequence analyses of the intact peptides and their cyanogen bromide digestion fragments with a gas-phase sequenator. The three human pancreas growth hormone-releasing factors contain 44 (hpGRF-44), 40 (hpGRF-40), and 37 (hpGRF-37) amino acids in identical sequences from their NH2 termini. High pressure liquid chromatography of the native peptides and their synthetic replicates showed that hpGRF-37 and hpGRF-40 possess free carboxyl termini while that of hpGRF-44 is amidated. The structure of hpGRF-44 was established as: Tyr-Ala-Asp-Ala-Ile-Phe-Thr-Asn-Ser-Tyr-Arg-Lys-Val-Leu-Gly-Gln-Leu-Ser-Ala Arg-Lys-Leu-Leu-Gln-Asp-Ile-Met-Ser-Arg-Gln-Gln-Gly-Glu-Ser-Asn-Gln-Glu-Arg-Gly Ala-Arg-Ala-Arg-Leu-NH2.

The hypophysiotropic peptide, corticotropin-releasing factor (CRF) was isolated from caprine hypothalamic median eminence tissue by means of acid extraction, immunoaffinity chromatography, gel filtration and two steps of reverse-phase high-performance liquid chromatography (RPLC). Amino acid sequence determination using a gas-phase sequencer established the primary structure of the first 39 residues from the NH 2 -terminus. The nature of the COOH-terminal dipeptide was elucidated by Staphylococcus aureus V8 protease digestion of the native peptide, dansylation of the digest and comparative RPLC studies with the dansylated dipeptides Ile-Ala-NH 2 , Ile-Ala-OH, Ala-Ile-NH 2 and Ala-Ile-OH. The complete structure of the peptide was established as: H-Ser-Gln-Glu-Pro-Pro-Ile-Ser-Leu-Asp-Leu-Thr-Phe-His-Leu-Leu-Arg-Glu-Val- Leu-Glu-Met-Thr-Lys-Ala-Asp-Gln-Leu-Ala-Gln-Gln-Ala-His-Ser-Asn-Arg-Lys-Leu- Leu-Asp-Ile-Ala-NH 2 , which is identical to that of ovine CRF.

A third melanotropin coding fragment named γ-MSH was discovered by Nakanishi et al (Nature 278 , 423–427 (1979)) in the cryptic region outside the portion coding for ACTH and β-LPH in the ACTH/β-LPH precursor mRNA isolated from the intermediate lobe of bovine pituitary. Four possible γ-MSH peptides derived from this coding fragment were synthesized by solid-phase methodology and their bioactivity determined in an in vitro MSH assay as well as the anterior pituitary primary culture assay. Relative to α-MSH, the melanotropic activities of Ac-γ1-MSH, γ1-MSH, γ2-MSH and γ3-MSH are 7.3 × 10−4, 3.3 × 10−5, 1.4 × 10−4 and 4.6 × 10−7 respectively. None of these γ-MSH peptides releases LH, FSH, PRL, GH and TSH in the pituitary culture medium at a dose as high as 100 ng per dish.

Peptides with high intrinsic activity to release growth hormone from pituitary cells in tissue cultures were isolated from two different human pancreatic tumors that had caused acromegaly. Homogeneous peptides were obtained after gel filtration and two steps of reverse-phase high-performance liquid chromatography. From one tumor a 44-residue peptide (human pancreas growth hormone releasing factor, hpGRF-44) was isolated, together with two shorter fragments of reduced bioactivity having 40 and 37 amino acid residues (hpGRF-40, hpGRF-37). In contrast, the other tumor contained only one form of GRF which proved to be identical to hpGRF-40. These hpGRFs are indistinguishable from partially purified preparations of hypothalamic growth hormone releasing factor of human, porcine and murine origins with respect to biological activity and are very similar in their physicochemical properties (molecular weight, retention behavior on reverse-phase HPLC, absence of sulfhydryl groups). One of the pancreatic tumors also contained two forms of immunoreactive somatostatin. One form, after isolation and partial microsequencing, was identified as somatostatin-14 with a structure identical to that of the peptide found in other species. The second form has tentatively been identified as somatostatin-28 on the basis of chromatographic behavior.

The release of GH induced by purified hypothalamic GRF or native or synthetic tumor-derived GRF is antagonized by the presence of CoCl2; it is simulated by 8Br .cAMP, IBMX, cholera toxin, forskolin, with identical maximal effects (Emax). Somatocrinin (GRF) stimulates the efflux of cAMP by the pituitary cells in parallel to the release of GH. Addition of either 8Br .cAMP, IBMX, cholera toxin or forskolin to a maximally stimulating dose of GRF does not increase the response which remains GRF-Emax. In contradistinction with these results PGE2 releases GH with a dose-response curve different from that of GRF, and the combination of PGE2 + GRF produces an Emax far greater than that due to either agonist alone; showing a true additivity. The name somatocrinin is proposed to replace the acronym GRF.

Summary This note describes the technical details of double antibody radio-immunoassays for the two peptides α- and β-endorphin and the specificity characteristics of the antisera. Antisera raised in rabbits to synthetic α-endorphin measure quantitatively α-endorphin; usable range is 50 pg to 5 ng. Their specific recognition site is toward the C-terminal region (Ser10-Thr16) of α-endorphin. Other antisera, raised in rabbits to synthetic β-endorphin measure quantitatively β-endorphin; usable range is 50 pg to 5 ng β-endorphin. Their specific recognition site is toward the C-terminal region (Asn20-His27) of β-endorphin; these antibodies recognize β-endorphin i.e. β-LPH-(61–91), β-lipotropin i.e. β-LPH-(1–91), and the precursor molecule to LPH-ACTH with molecular weight ca. 31 × 103 (31K-precursor). Neither antiserum binds enkephalins.

A specific radioimmunoassay for ..gamma..-melanocyte stimulating hormone-like peptides was developed. An antiserum raised in rabbit to synthetic bovine ..gamma../sub 3/-MSH, one of the possible ..gamma..-MSH peptides, specifically recognizes the portion between His/sup 5/ and Arg/sup 14/ of ..gamma../sub 3/-MSH without significant cross-reaction with other synthetic ..gamma..-MSH-like peptides, ..cap alpha..-, ..beta..-MSH, adrenocorticotropin, and ..beta..-endorphin. The usable range of this RIA is 10 pg to 600 pg of synthetic ..gamma../sub 3/-MSH. Three immunoreactive ..gamma..-MSH peaks were thus found in gel permeation chromatography of the whole bovine pituitary extract.

In view of striking similarities between TGF-beta and inhibin, we investigated the possibility that TGF-beta might modulate pituitary hormone release in vitro. Long term incubations of beta transforming growth factor (TGF-beta) with rat anterior pituitary cells for 48 hr stimulates the basal secretion of FSH in a dose-dependent manner. The secretion of LH, TSH, GH, ACTH and PRL is not modified by TGF-beta. The minimal effective concentration of TGF-beta is 10 pg/ml (less than 500 attomolar) and is dose dependent over a range from 1 pg to 10 ng/ml. Treatment of cells with TGF-beta for short incubation times (4 hr) in assays similar to that used for hypophysial releasing factors is not effective, indicating that TGF-beta acts through a cellular mechanism distinct from that of LRF. Inhibin-A, recently characterized on the basis of its capacity to specifically inhibit the secretion of FSH in the 48 hr bioassay system inhibits the stimulatory effect of TGF-beta on FSH-release. Analyses of the dose response curves indicate that the interaction occurs in a typical non-competitive manner. The results suggest that a TGF-beta-like molecule, present in follicular fluid, may be responsible for the FSH-releasing activity ("anti-inhibin" activity) observed by us and others during the process of isolating inhibin from follicular fluids. They also suggest an important role for inhibin and the TGF-beta related molecules in modulating pituitary gonadotropin release.

The synthesis by solid-phase methodology of two glycosylated analogs of somatostatin [Glc-Asn 5 ]-SS and [NAcGlc-Asn 5 ]-SS is described. These two analogs have been biologically tested on the secretion of pituitary growth hormone, pancreatic glucagon and insulin. The results show that glycosylation of somatostatin on the Asn 5 residue decreases by a hundred fold the inhibition activity on GH release when tested in vitro . In vivo , since the activity is similar to somatostatin the carbohydrates are probably removed by some enzymatic reaction and thus liberate the full activity of somatostatin.

Immunoreactive-calcitonin was observed in all cells of the intermediate lobe of the rat pituitary gland. It may thus be a fragment of the ≥31, 000 daltons precursor molecule of adrenocorticotropin and β-lipotropin.

A 41 amino acid peptide with high intrinsic corticotropin-releasing activity was isolated from 1000 bovine hypothalami by means of immunoaffinity chromatography, gel filtration, and two steps of reverse phase HPLC. The primary structure of the amino terminal 39 amino acids was characterized by gas phase sequence analysis. The sequence of the amidated car☐yl terminal dipeptide was established by digestion of the intact natural product with Staphylococcus aureus V8 protease, dansylation of the digest and comparative reverse phase liquid chromatography studies with the synthetic dansylated dipeptides Ile-Ala-NH 2 , Ile-Ala-OH, Ala-Ile-NH 2 and Ala-Ile-OH. The complete structure of the bovine corticotropin-releasing factor was established as: Ser-Gln-Glu-Pro-Pro-Ile-Ser-Leu-Asp-Leu-Thr-Phe-His-Leu-Leu-Arg-Glu-Val-Leu- Glu-Met-Thr-Lys-Ala-Asp-Gln-Leu-Ala-Gln-Gln-Ala-His-Asn-Asn-Arg-Lys-Leu-Leu- Asp-Ile-Ala-NH 2 using approximately 650 pmol of material.

Extraction of bovine pituitaries in the presence of enzyme inhibitors (2 mM PMSF, 2 mM sodium tetrathionate, 15 microM pepstatin A, and 1 mM EDTA) resulted in the isolation of two distinct forms of basic fibroblast growth factor. Partial characterization of both molecules showed one form to be identical to basic FGF(1-146) which has already been reported by our laboratory. The second form was estimated by SDS-PAGE to have a molecular weight of 17,000 Daltons which is slightly larger than that of basic FGF(1-146). Amino acid analysis shows the presence of 8 new residues more than basic FGF(1-146) which accounts for the difference in molecular weight. Gas-phase sequencing of this molecule indicated that it bears a blocked amino terminus. Furthermore, this higher molecular weight form of basic FGF did not show immunoreactivity with antibodies specific for the amino terminus of basic FGF(1-146) but cross reacted with antibodies generated against midportion fragments of basic FGF(1-146), indicating that the molecule is amino terminally extended. Like basic FGF(1-146), the molecule is a potent mitogenic factor for vascular endothelial cells. Taken together these results demonstrate the existence of a precursor form of basic FGF which is extended by 8 residues at the amino terminus with the first residue being blocked.

A radioimmunoassay (RIA) was developed to measure fibroblast growth factor (FGF) using antiserum generated against a synthetic replicate of [Tyr 10 ]FGF(1–10). The antisera, previously shown to be capable of inhibiting the biological action of FGF on bovine aortic arch endothelial cells in vitro [1], are highly specific for the amino-terminus of FGF. In the RIA, the antisera recognize the decapeptide antigen [Tyr 10 ]FGF(1–10) and the intact mitogen on an equimolar basis and show less than 0.01% cross-reactivity with N -acetyl-[Tyr 10 ]FGF(1–10). Bovine adenohypophysial cells maintained in primary monolayer culture release and ir-FGF which is indistinguishable from the intact mitogen in as much as it is retained on heparin-Sepharose affinity columns and shows a dose-dependent and parallel displacement in RIA. The release of ir-FGF by the bovine adenohypophysis can be increased with forskolin (10 −5 M) or KCl (50 mM). Preincubation of pituitary cells with 17β-estradiol has no measurable effects on basal ir-FGF, but increases the release after KCl treatment 2–3-fold. These results show that ir-FGF can be released by the bovine adenohypophysis in vitro and lend credence to the hypothesis that FGF plays a physiological role in the homeostatic mechanisms regulating mesoderm-derived cell growth.

The action of cysteamine on anterior pituitary hormone secretion was studied in vivo using conscious, freely moving male rats and in vitro using anterior pituitary cells in monolayer culture. Administration of 500 μg cysteamine into the lateral cerebral ventricles of normal rats caused the complete inhibition of pulsatile GH secretion for a minimum of 6 h. This treatment also significantly decreased plasma concentrations of LH for at least 6 h in orchiectomized rats, TSH in short-term (0.5 month) thyroidectomized rats, and PRL in long-term (6 months) thyroidectomized rats. The in vivo stimulation of GH, LH, TSH and PRL with their respective releasing hormones 60 min after administration of cysteamine was not different from the response observed in rats pretreated with saline except for PRL where cysteamine pretreatment significantly inhibited the expected PRL increase. In vitro, 1 mM cysteamine decreased basal and TRH stimulated PRL release while not affecting basal or stimulated GH, LH, TSH and ACTH secretion. These data demonstrate the dramatic and wide-ranging effects of cysteamine on anterior pituitary hormone secretion. This action appears to be mediated through hypothalamic pathways for GH, LH and TSH and through a pituitary pathway for PRL.

Rapid and prolonged analgesia was obtained in all of 14 obstetric patients who received synthetic human β-endorphin intrathecally at the time of delivery. Normal uterine contractions were maintained and all women were fully conscious and highly cooperative in the delivery process. No depression of respiration rate, cardiovascular or central nervous system was observed in any of the patients. Conditions of the infants evaluated by Apgar scoring were excellent. β-endorphin must be administered intrathecally because it does not cross the blood-brain barrier; for the same reason, β-endorphin cannot enter the central nervous system of the fetus as do opiates or other drugs commonly used as anesthetics or analgesics at the time of delivery.

(1) α-, β-, γ-endorphins have primary structures respectively identical to those of β-lipotropin subunits [61–76], [61–91], [61–77]. (2) These peptides exhibit opiate-like activity in several assay systems. (3) A series of analogs of the naturally occurring peptides has been prepared and their biologic activities compared. (4) Each of the endorphins produces a discrete set of profound behavioral changes when injected into the ventricular system of the brain of rats. (5) The hypothesis is advanced that alterations in the homeostasis of the β-lipotropinendorphin system may be causally involved in the mental diseases of man.

Five NH 2 -terminal extension analogs of methionine-enkephalin have been synthesized by solid phase methodology and their morphinomimetic activity determined in the guinea pig ileum-myenteric plexus preparation. Relative to methionine-enkephalin which corresponds to β-LPH-(61–65) and arbitrarily assigned a potency value of 100, β-LPH-(60–65), β-LPH-(59–65), β-LPH-(58–65), β-LPH-(57–65) and β-LPH-(56–65) have potencies of 90, 7, 4.4, 0.3 and 0.11 respectively.

Somatocrinin, a 44 amino acid peptide with potent growth hormone (GH) releasing activity in anesthetized rats, was tested in conscious freely-moving rats. When high doses of 1 to 10 μg were administered (iv) at random times between spontaneous GH pulses, the responses were inconsistent. When similar doses were tested under identical conditions but in rats pretreated with antibodies against somatostatin, all animals demonstrated a marked and immediate increase in plasma GH of 5 to 10 fold. Similarly, a 1 μg dose of somatocrinin was also ineffective in increasing plasma GH when administered to rats subjected to a 72 h fast, a paradigm known to enhance endogenous somatostatin secretion. However, plasma GH increased over 20 fold if rats were pretreated with antibodies against somatostatin. These results demonstrate the dynamic and opposite roles exerted by somatocrinin and somatostatin in regulating GH secretion.

Peptides with high intrinsic growth hormone releasing activity (growth hormone-releasing factor, GRF) were isolated from 2100 ovine and 2600 caprine (goat) hypothalami by means of acid extraction, immunoaffinity chromatography, gel filtration and reverse phase HPLC. Structural characterization of the 44 amino acid ovine peptide by gas-liquid phase sequencing and peptide mapping established its primary structure as Tyr-Ala-Asp-Ala-Ile-Phe-Thr-Asn-Ser -Tyr-Arg-Lys-Ile-Leu-Gly-Gln-Leu-Ser-Ala-Arg-Lys-Leu-Leu-Gln-Asp-Ile-Met -Asn -Arg-Gln-Gln-GLy-Glu-Arg-Asn-Gln-Glu-Gln-Gly-Ala-Lys-Val-Arg-Leu-NH2. Caprine GRF was found to possess the same sequence except for the replacement of the isoleucine residue in position 13 with valine and thus is identical to bovine GRF.

The tetrapeptide pGlu-Tyr-Arg-Trp-NH2 was synthesized by solid phase methodology; its characterization by nuclear magnetic resonance and mass spectrometry is demonstrated. Contrary to earlier claims in the literature, pGlu-Tyr-Arg-Trp-NH2 stimulates the release of both luteinizing hormone and follicle stimulating hormone. Its specific activity for the release of either gonadotropin when compared to the decapeptide LRF (luteinizing hormone releasing factor) is ca. 1100,000th that of LRF.

Summary A purification sequence for the ovine hypothalamic LH-releasing factor is reported. From 300,000 sheep brains, ca. 200 μg have been obtained of a material containing ⪯ 56% peptide, of which 94% can be accounted for by the amino acids His 1, Arg 1, Ser 1, Glu 1, Pro 1, Gly 2, Leu 1, Tyr 1, the N-terminal being pyroGlu (2-pyrrolidone-5-carboxylic acid, PCA). This material stimulates release of LH in vivo and in vitro (≥0.5 ng/ml); it also stimulates release of FSH concomitantly with LH.

Specific binding of 3H-labelled TRF to mouse thyrotropic tumor membrane preparations and to intact pituitary cells is described. The binding is saturable with respect to 3H-TRF under the conditions used and stoichiometrically competible by unlabelled TRF providing the basis for a workable in vitro assay for this releasing factor. Neither luteinizing hormone releasing factor (LRF) nor a biologically inactive dipeptide derivative of TRF compete for 3H-TRF binding. The dissociation constant for 3H-TRF binding to the tumor plasma membrane fraction is ca. 4 × 10−8M at 0°C.

Discrete cells containing neurotensin, as shown by immunofluorescence, have been observed in the lower portion of the dog ileum. This implies that neurotensin may be synthesized in the small intestine and may be involved in local regulation of intestinal functions. Neurotensin is a peptide characterized originally in the hypothalamus.

Profound and long-lasting analgesia (mean duration of pain relief 33.4 h, range 22.5--73.5 h) was produced by intrathecal administration of 3 mg synthetic beta-endorphin in all of 14 patients with intractable pain due to disseminated cancer. No respiratory depression, hypotension, hypothermia, or catatonia was observed.

Immunoassayable β-endorphin is present in samples of amniotic fluid obtained in the last two weeks of pregnancy. Average concentration is approximately 300 pg. per milliliter. In all cases of suspected or recognized fetal distress, striking elevations of the β-endorphin concentrations (twofold to 20-fold) were observed. The degree of elevation correlated with the degree of fetal distress.