The distribution of somatostatin-like immunoreactive (SS-LI) material and its colocalization with glutamic acid decarboxylase (GAD)-like immunoreactivity were studied in the rat hippocampus and dentate gyrus neurons using immunohistochemistry. In the dentate gyrus and CA1 region, SS-LI perikarya were concentrated in the hilus and in the stratum oriens, respectively, whereas immunoreactive cell bodies were rarely seen in other layers. Approximately half of the SS-LI neurons of the CA3 region were situated in the stratum oriens, the other half being scattered in strata pyramidale, lucidum and radiatum. About 90% of SS-LI neurons were also GAD-like immunoreactive, whereas about 14% of GAD-like immunoreactive (GAD-LI) neurons were SS-like immunoreactive. The percentage of GAD-LI neurons which were also immunoreactive for SS varied from one layer to the other. This percentage was about 30% in the hilus of the dentate gyrus and in the stratum oriens of the CA1 and CA3 regions; it was 5-10% in the strata pyramidale, lucidum and radiatum of the CA3 region and reached only 2% in the granule cell layer and molecular layer of the dentate gyrus and in the stratum pyramidale and stratum radiatum in the CA1 region. These observations indicate that the majority of SS-LI neurons in the rat hippocampal formation are a subpopulation of GABAergic neurons.

Approximately one half of the neurons in the lumbar dorsal root ganglion of adult rats display high-affinity receptors for nerve growth factor (NGF). To ascertain which types of sensory neurons are potentially responsive to NGF, adjacent cryostat sections of rat dorsal root ganglia were processed either for NGF-receptor using radioautography or by one of four histochemical procedures. Histograms of the densities of neuronal labelling by radioiodinated NGF were examined for subpopulations of lumbar sensory neurons with thiamine monophosphatase enzyme activity or with immunoreactivity for calcitonin gene-related peptide (CGRP), substance P, or somatostatin. Virtually all neurons with strong CGRP immunoreactivity had high-affinity NGF binding sites, although some neurons with faintly positive CGRP immunoreactivity lacked such NGF binding. A subpopulation of large neurons, approximately 5% of the total, had dense labelling by 125I-NGF but were not stained by this immunohistochemical technique for CGRP. Of the three major populations of small neurons those with substance P immunoreactivity were consistently and heavily labelled by radioiodinated NGF whereas those with somatostatin immunoreactivity or thiamine monophosphatase activity were not specifically labelled by radioautography. For these primary sensory neurons in mature rats the genes for substance P and CGRP seem to be strongly expressed only in neurons capable of responding to NGF. On the other hand, neurons containing somatostatin and thiamine monophosphatase invariably lack high-affinity NGF receptors.

No abstract

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. We have further investigated this hypothesis by studying the anatomical organization of GRF neurones in rat hypothalamus, using an antibody raised against the recently characterized rat hypothalamic GRF in normal animals and in animals neonatally treated with monosodium glutamate (MSG), a treatment which results in the selective destruction of arcuate nucleus neurones. We present here the results which show that GRF-producing neurones are located mainly in the arcuate nucleus of rats. MSG treatment results in the complete loss of GRF-immunoreactive cell bodies within this nucleus and provokes a selective disappearance of GRF-immunoreactive fibres in the median eminence. These results show that the arcuate nucleus is the origin of the GRF-containing fibres that project to the median eminence and establish the MSG-treated rat as an in vivo model for studying growth hormone secretion in the absence of neurohumoral GRF.

Senile dementia of the Alzheimer's type can be diagnosed with certainty only by examining neurofibrillary tangles and neuritic plaques under the microscope. Recently, it has been suggested that the condition is linked to specific neurotransmitter systems, with a decline of cortical acetylcholine, choline acetyltransferase, cholinergic neurones projecting to the cortex, cortical noradrenaline content, locus coeruleus neurones and cortical somatostatic content. Using immunocytochemical methods, we here report that somatostatin-immunoreactive processes are present in neuritic plaques in human Alzheimer's specimens. These data, as well as other reports of non-cholinergic changes, strongly imply that Alzheimer's disease cannot be linked exclusively to cortical cholinergic elements, as proposed previously. Rather, our data on plaque and somatostatin co-localization and distribution patterns suggest that Alzheimer's neuropathology may involve primarily the loss of selective cortical neurones that are targets of the implicated transmitter systems and that plaque formation may result from the degeneration of presynaptic and postsynaptic neurites of large projection neurones in layers III and V. Given the neurochemically heterogeneous input to these cells, it is not surprising that several neurotransmitter systems, one of which is somatostatin, are implicated in the pathology of Alzheimer's disease.

Cleavage of the peptide bonds of preprosomatostatin at basic residues near the carboxyl terminus yields somatostatin-14, somatostatin-28, and somatostatin-28 (1-12). However, little is known about the molecular forms derived from the amino terminal portion of the precursor, even though this part of the prohormone is highly conserved through evolution. By using an antibody against the amino terminus of prosomatostatin, a decapeptide with the structure Ala-Pro-Ser-Asp-Pro-Arg-Leu-Arg-Gln-Phe, corresponding to preprosomatostatin (25-34), was isolated from the endocrine portion of the rat stomach, the gastric antrum. The antral decapeptide may represent a bioactive product generated from prosomatostatin after a monobasic cleavage similar to that involved in the formation of somatostatin-28. In fact, a monobasic cleavage requires two basic residues and a domain containing nonpolar amino acids such as alanine or leucine, or both.

The primary structure of the NH2terminally extended somatostatins isolated from ovine hypothalamic extracts, one containing 28 residues and the other 25, has been determined. The structure of somatostatin-28 is Ser-AlaAsn-Ser-Asn-Pro-Ala-Met-Ala-Pro-Arg-Glu-Arg-Lys-Ala-GlyCys-Lys-Asn-Phe-Phe-Trp-Lys-Thr-Phe-Thr-Ser-Cts-OH; the shorter one, somatostatin-25, has the same sequence as somatostatin-28 except that the first three NH2-terminal residues are deleted. The two peptides as isolated were found to be oxidized at the methionine residue to the methionine sulfoxide. Their structures were established by subjecting the native peptides to direct sequence analysis in a Beckman 890C sequencer and identifying the released phenylthiohydantoin derivatives by high-performance liquid chromatography. Their structures were confirmed by trypsin digestion and isolation of all the tryptic peptides, followed by amino acid analysis of the tryptic fragments. Moreover, some of the tryptic peptides were matched with their respective synthetic replicates on high-performance liquid chromatography.The tetradecapeptide somatostatin was first isolated and characterized from ovine hypothalamus (1) on the basis of its ability to inhibit the secretion of growth hormone from primary cultures of dispersed rat anterior pituitary cells. The identical tetradecapeptide was subsequently isolated and characterized from porcine hypothalamus (2) as well as pigeon (3) and anglerfish (4) pancreases. In addition, several laboratories (2, 3, 5-8) have observed, in the hypothalamic and pancreatic extracts, bioactive and immunoreactive somatostatin-like substances having molecular weights larger than the tetradecapeptide. Pradayrol et al. (9) reported the isolation (from porcine intestinal extracts) and primary structure of an NH2-terminally extended somatostatin containing 28 amino acids which was designated SS-28. We have also isolated two bioactive somatostatins containing 28 and 25 residues from ovine hypothalamic extracts. The primary structure of the larger ovine hypothalamic somatostatin is Ser-Ala-Asn-Ser-Asn-Pro-Ala-Met-AlaPro-Arg-Glu-Arg-Lys-Ala-Gly-Cys-Lys-Asn-Phe -Phe -TrpLys-Thr-Phe-Thr-Ser-Cys-OH, which is identical to the structure of the intestinal SS-28 isolated by Pradayrol et al. (9). The shorter ovine hypothalamic peptide, SS-25, has the same structure as SS-28 except that the first three NH2-terminal residues are deleted.We report here in detail the determination of the primary structure of the two larger hypothalamic somatostatins.The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact. 6827MATERIALS AND METHODS The starting material for the isolation work was the combined Sephadex G-25 excluded fractions from extracts of 350,000 ovine hypothalami used in our purification of the thyrotropin-releasing factor (10). Details of the isolation procedure will be published elsewher...

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). However, a 44-amino-acid amidated peptide having the physiological properties of GRF as well as chemical similarities was recently isolated from a human pancreatic tumour that had caused acromegaly. Two shorter biologically active fragments of 40 and 37 residues were also isolated. The synthetic replicates of these human pancreas GRF (hpGRF) peptides specifically stimulate GH release in vitro and in vivo. Assuming similarity or identity between the putative hypothalamic GRF and the tumour-derived hpGRF, we have used immunohistochemistry to search for hpGRF-like immunoreactivity in the brain. We report here that antisera against the hpGRF1-40 peptide specifically stain neuronal cell bodies in the arcuate nucleus of the primate hypothalamus, with fibres projecting to the median eminence and ending in contact with portal vessels. This topography is characteristic of a neuronal system elaborating a releasing factor. These results provide evidence that hypothalamic GRF is very similar, if not identical, to hpGRF.