Subcellular distribution of somatostatin-14, somatostatin-28 and somatostatin-28 (1-12) in rat brain cortex and comparisons of their respective binding sites in brain and pituitary.

Subcellular distribution and binding characteristics of the three endogenous peptides somatostatin-14 (SRIF-14), somatostatin-28 (SRIF-28) and somatostatin-28(1-12) (SRIF-28(1-12] derived from preprosomatostatin were investigated in the rat brain cortex. The three peptides are predominantly recovered from a crude mitochondrial pellet (P2), containing the pinched off nerve endings. Specific high affinity binding sites for 125I-N-Tyr-SRIF-14 and 125I-N-Tyr-SRIF-28 are present on pituitary and brain membranes.

Isolation and characterization of the bovine hypothalamic corticotropin-releasing factor.

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 carboxyl 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-NH2, Ile-Ala-OH, Ala-Ile-NH2 and Ala-Ile-OH.

The distribution and morphological characteristics of the intracortical VIP-positive cell: an immunohistochemical analysis.

Using a sensitive immunohistochemical procedure, we have undertaken a detailed morphological characterization of the vasoactive intestinal peptide-positive (VIP-positive) neuron in the cerebral cortex of the rat. VIP-positive neurons are present in all regions of cortex, and are usually strongly bipolar, possessing long, radially directed processes with very limited branching in the tangential plane. The most extensive dendritic branching occurs in layers I and deep IV-superficial V, and the density of axonal varicosities is highest in layers II-IV.

Evidence for selective release of somatostatin-14 and somatostatin-28(1-12) from rat hypothalamus.

Cysteamine administration to rats results in a marked depletion of hypothalamic somatostatin-14 (SS14) and a decrease of the potassium-evoked in vitro release of SS14 without a significant change in the content or release of somatostatin-28(1-12)-like immunoreactivity (SS28(1-12)-L1). Furthermore, cysteamine enhances the spontaneous release and markedly potentiates the potassium-evoked release of SS14 in the in vitro slice preparation. However, in vitro-administered cysteamine does not alter the spontaneous or potassium-evoked release of SS28(1-12)-LI.

Specific depletion of immunoreactive growth hormone-releasing factor by monosodium glutamate in rat median eminence.

A potent and specific growth hormone-releasing factor (GRF) was recently isolated and characterized from a human islet cell tumour of the pancreas that caused acromegaly. Antibodies raised against the synthetic replicate of this peptide have allowed the immunohistochemical identification of GRF-producing neurones within the primate central nervous system. Such neurones are found mainly in the arcuate nucleus in human and monkey hypothalamus, suggesting that this nucleus is a primary source of GRF.

Somatostatin-28(1-12)-like immunoreactive substance is secreted into hypophysial portal vessel blood in the rat.

Somatostatin-28(1-12)-like immunoreactivity was measured in extracts of rat hypophysial portal blood and peripheral blood. The concentration of somatostatin-28(1-12) was higher in portal than in peripheral extracts, and its release into portal vessel blood was increased 4- to 5-fold by electrical stimulation of the median eminence. These results show that somatostatin-28(1-12) may be a physiological neurohormone and/or neurotransmitter.

Pancreatic somatostatinoma: abundance of somatostatin-28(1-12)-like immunoreactivity in tumor and plasma.

Unlabelled: In the present study we characterized and compared the relative amounts of the different molecular forms of somatostatin-14 like immunoreactivity (S-14 LI) and of somatostatin-28(1-12) like immunoreactivity (S-28(1-12) LI) in extracts of tumor and peripheral plasma of a patient with a pancreatic somatostatinoma. Tissue and plasma were chromatographed on Sephadex G-50 columns equilibrated with 6 M urea. Immunoreactivity in the eluting fractions was assayed with two separate, region specific RIAs using antibodies R149 (S-14 LI) and S309 (S-28(1-12)LI).

Inhibitory effects of cysteamine on neuroendocrine function.

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 micrograms 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 rat, TSH in short-term (0.

Effects of cysteamine on pro-somatostatin related peptides.

Intracerebroventricular (icv) injection of cysteamine to rats produced a marked depletion of somatostatin-14-like immunoreactivity (LI) in all rat brain regions examined. The somatostatin-28 (SS28)-LI and SS28(1-12)-LI were generally not altered by the cysteamine treatment. Following subcutaneous injection of the drug similar depletions of hypothalamic SS14-LI was observed with no change in SS28-LI nor SS28(1-12)-LI.

Immunohistochemical evidence that brain enkephalins arise from a precursor similar to adrenal preproenkephalin.

We have examined rat brains by immunohistochemistry with an antiserum raised against BAM22P, a synthetic fragment of the adrenomedullary precursor of Leu- and Met-enkephalin, in order to determine whether preproenkephalin derived peptides are detectable in the central nervous system. BAM22P antiserum stained fibers, nerve endings and cell bodies in many areas of the brain, especially the striatum, the septum and the hypothalamus. Comparative topographical studies showed that neurons recognized by anti-BAM22P overlapped structures found immunoreactive for enkephalins in the present study.

Immunohistochemical distribution of pro-somatostatin-related peptides in cerebral cortex.

Mammalian brain contains 3 peptides related to the pro-somatostatin molecule: somatostatin-14 (SS14), the form originally identified from hypothalamic extracts, somatostatin-28 (SS28) and somatostatin 28 (1-12) (SS28 (1-12)). By using antibodies which selectively recognize one or more of these 3 somatostatin-related peptides, we have characterized their immunohistochemical distribution in neocortex. These somatostatin-related peptides have a specific laminar distribution in cortex and are differentially distributed such that SS28 is largely restricted to cell bodies, whereas SS28 (1-12) is preferentially localized in neuronal processes and terminals in a density which far exceeds that revealed by SS-14 immunoreactivity.

Immunohistochemical detection of growth hormone-releasing factor in brain.

The concept of a hypothalamic neurohumoral control for anterior pituitary secretion postulates the existence of a growth hormone-releasing factor (GRF) of neuronal origin that stimulates the pituitary gland to release growth hormone (GH). Such a compound has not yet been isolated and characterized from the brain, although there is extensive physiological and biochemical evidence for its existence (reviewed in ref. 2).

Release of somatostatin-28(1-12) from rat hypothalamus in vitro.

Following the discovery of the growth hormone release-inhibiting factor somatostatin from extracts of ovine hypothalamus, an N-terminally extended somatostatin of 28 amino acids has been identified in mammalian tissue. The original peptide, somatostatin-14 (SS14), corresponds to the C-terminus of somatostatin-28 (SS28). Both SS28 and SS14 have biological activity, occur in several rat brain regions, are present in cell bodies and nerve terminals and can be released in vitro upon depolarization in a calcium-dependent manner.

Pituitary adenomas that caused Cushing's disease or Nelson's syndrome are not responsive to ovine corticotropin-releasing factor in vitro.

The response of pituitary adenomas obtained surgically from patients with Cushing's disease of Nelson's syndrome to synthetic ovine corticotropin-releasing factor (CRF), vasopressins, somatostatin-28, dexamethasone, 3-isobutylmethylxanthine or high [K+] was examined in vitro by measuring the amount of pro-opiomelanocortin (POMC)-derived peptides secreted into the culture medium. CRF did not stimulate the secretion of adrenocorticotropin-, beta-endorphin-, or gamma 3-melanotropin-like peptides from the pituitary adenomas at concentrations ranging from 1 x 10(-13) M to 1 x 10(-7) M whereas vasopressins, 3-isobutyrl-methylxanthine and high [K+] increased, while somatostatin-28 and dexamethasone suppressed, the secretion of these POMC-derived peptides. These findings suggest that either the pituitary ACTH-producing tumors have lost their receptors to CRF or their post-receptor mechanism to CRF is not functional.

Anaerobic degradation of cyanuric Acid, cysteine, and atrazine by a facultative anaerobic bacterium.

A facultative anaerobic bacterium that rapidly degrades cyanuric acid (CA) was isolated from the sediment of a stream that received industrial wastewater effluent. CA decomposition was measured throughout the growth cycle by using a high-performance liquid chromatography assay, and the concomitant production of ammonia was also measured. The bacterium used CA or cysteine as a major, if not the sole, carbon and energy source under anaerobic, but not aerobic, conditions in a defined medium.

Immunocytochemical localization of prosomatostatin fragments in maturing and mature secretory granules of pancreatic and gastrointestinal D cells.

Pancreatic and gastrointestinal D cells were examined by immunocytochemistry using antisera against somatostatin-28 (SS28) and its NH2-terminal fragment SS28-(1-12), followed by the staphylococcal protein A-gold (pAg) complex. In pancreatic and gastric D cells incubated with antiserum against SS28-(1-12) the gold particles produced intense staining of the mature secretory granules but weaker staining of the immature granules associated with the Golgi area, whereas after SS28 antiserum treatment the particles accumulated selectively over the population of immature secretory granules. In intestinal D cells not only SS28-(1-12) but also SS28 antiserum produced an intense gold staining over the mature delta granules.

Immunohistochemical distribution of pro-somatostatin-related peptides in hippocampus.

By using immune sera which recognize one or more of the 3 peptides, somatostatin-14 (SS14), somatostatin-28 (SS28) and somatostatin-28(1-12) (SS28(1-12)) we have characterized their immunohistochemical distribution in the hippocampal formation. There exist at least two independent neuronal systems containing pro-somatostatin-related peptides: an intrinsic system of cells in the polymorphic layers which branch locally, and a dense terminal field in the molecular layer of the dentate gyrus that may constitute a portion of the entorhinal-dentate projection. In addition, SS28 is the dominant form present in cell bodies, whereas SS28(1-12) is preferentially localized in neuronal processes and terminals.

Somatostatin-28(1-12)-like immunoreactivity in the rat.

The distribution of the newly characterized peptide somatostatin-28(1-12) was determined in rat tissues using a radioimmunoassay in which the immune plasma is directed against the C-terminus of the dodecapeptide. In the central nervous system, somatostatin-28(1-12)-like immunoreactivity is highly concentrated in the hypothalamus and the amygdala. In the digestive system, the highest levels of immunoreactivity are found in the stomach, the pancreas and the colon.

Molecular forms of the putative enkephalin precursor BAM-12P in bovine adrenal, pituitary, and hypothalamus.

A highly specific radioimmunoassay for one of the putative adrenomedullary [Met]enkephalin precursors, BAM-12P (Tyr-Gly-Gly-Phe-Met-Arg-Arg-Val-Gly-Arg-Pro-Glu-OH), has been developed. The BAM-12P antibodies recognize the COOH-terminal fragment of the peptide from Arg7 to Glu12 and do not crossreact with [Met5]- or [Leu5]enkephalin or any of their COOH-terminal lysine or arginine extended analogs. Specificity for the COOH-terminal Glu-OH is suggested by the 100% crossreactivity with BAM-12P5-12 and 0.

Presence of somatostatin-28-(1-12) in hypothalamus and pancreas.

Acid extracts from rat pancreas and hypothalamus were analyzed for the presence of the antigenic determinant corresponding to the NH2 terminus of somatostatin-28 (SS28), using an antiserum directed against amino acids 1 to less than or equal to 11 of the SS28 molecule. On gel permeation chromatography the majority of the immunoreactive material from each tissue extract eluted in one zone compatible with a peptide of 1250 daltons. Purification of this immunoreactive material by reverse-phase HPLC and cation-exchange chromatography yielded two immunoreactive peptides from each tissue extract.

Isolation and characterization of rat hypothalamic somatostatin-14.

A peptide with somatostatin-like immuno- and bioactivity has been isolated from 1165 rat hypothalami by using antisomatostatin affinity chromatography, gel filtration and reverse-phase high-performance liquid chromatography. The isolated peptide is indistinguishable from synthetic somatostatin-14 with respect to chromatographic properties and amino acid composition. We therefore propose that rat hypothalamic somatostatin-14 is identical in structure to somatostatin-14 found in other species.