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Several peptides are generated from prosomatostatin (proSS) in addition to somatostatin-14 (SS-14) and somatostatin-28 (SS-28). These are SS-28(1–12), proSS(1–76) and, as shown more recently, proSS(1–63) and antrin. Important variations in the proportion of these molecular forms are seen among different tissues and among different species. Processing of the precursor in the human brain yields minimal quantities of SS-28(1–12) and high levels of proSS(1–76), namely in cortical areas and in the bed nucleus of the stria terminalis. This is in contrast with findings in rat brain, where SS-28(1–12) is a predominant molecular form. Antrin, which corresponds to proSS(1–10), reaches its highest concentration in the antral portion of the stomach (117 ± 13 pmol/g wet weight), where it is found in secretory granules of delta cells. We observed an inverse relationship between levels of antrin and proSS(1–63) after chromatography of various tissue extracts. This suggests a precursor-product relationship between these two peptides.

The biochemical characterization and anatomical distribution of somatostatin binding sites were examined in the brain of the frog Rana ridibunda, and the distribution of the receptors was compared with the location of somatostatin immunoreactive neurons. The pharmacological profile of somatostatin receptors was determined in the frog brain by means of an iodinated superagonist of somatostatin, [125I-Tyr0,DTrp8]S-14. Membrane-enriched preparations from frog brain homogenates were shown to contain high-affinity receptors (KD = 0.78 ± 0.34 nM; Bmax = 103 + 12.7 fmoles /mg protein) with pharmacological specificity for [DTrp] substituted S14 and S28 analogs. The distribution of somatostatin-binding sites was studied by autoradiography on coronal sections of frog brain. Various densities of somatostatin receptors were detected in discrete areas of the brain. The highest concentration of binding sites was observed in the olfactory bulb, in the pallium, and in the superficial tectum. Moderate binding was observed in the striatum, amygdaloid complex, preoptic area, and cerebellum. Immunocytochemical studies of the distribution of somatostatin-28 (S28) related peptides were also conducted in the frog brain. Two antisera that recognize distinct epitopes of the somatostatin molecule have been used for immunohistochemical mapping of the peptide. Antiserum SS9 recognizes both S28 and somatostatin-14 (S14) and allowed the labelling of perikarya. Antiserum S320 recognizes the N-terminal fragment (1–12) resulting from enzymatic cleavage of S28. This latter antiserum, which does not cross-react with S28, stained mainly neuronal processes. At the infundibular level, however, both antisera stained cell bodies and fibers. Immunoreactive somatostatin-related peptides were detected in many areas of the frog brain. In the diencephalon, a heavy accumulation of perikarya and fibers was seen in the preoptic nucleus, the dorsal and ventral infundibular nuclei, and the median eminence. Immunoreactive perikarya were also observed in the telencephalon, especially in the pallium and in thalamic nuclei. Immunostained processes were detected in many telencephalic areas and in the tectum. There was good correlation between the distribution of somatostatin-immunoreactive elements and the location of somatostatin-binding sites in several areas of the brain, in particular in the median pallium, the tectum, and the interpeduncular nucleus. In contrast, mismatching was observed in the olfactory bulb, lateral pallium, and the cerebellum (which contained moderate to high levels of binding sites but virtually no somatostatin-immunoreactive fibers) and the hypothalamic areas (preoptic area and infundibular nuclei), in which a low concentration of binding sites but a high density of immunoreactive perikarya and fibers were detected. The present data provide the first pharmacological characterization and anatomical distribution of somatostatin-binding sites in the brain of a non-mammalian vertebrate species. The present results suggest that many of the major features of somatostatinergic systems described in mammals (e.g., the wide distribution of somatostatinergic neurons and somatostatin-binding sites; pharmacological characteristics of somatostatin receptors) had already arisen in tetrapod ancestors. Such findings suggest that both neuroendocrine and extrahypothalamic somatostatinergic systems have important functions throughout the vertebrate phylum.

A deficiency in somatostatin is the most consistently described neurochemical alteration in Alzheimer's disease (AD) attributable to intrinsic cortical neurons. Somatostatin-28 (SOM-28), an N-terminal-extended form of somatostatin, can be cleaved to form somatostatin-28(1-12)(SOM-28(1-12) ) and somatostatin-14 (SOM-14). We have measured concentrations of SOM-28(1-12)-like immunoreactivity in 8 cortical regions from 12 patients with AD and 13 controls. Significant reductions (P less than 0.001) were found in all cortical regions examined with the largest decrease in temporal lobe. Reductions were significantly correlated with decreases in somatostatin-14-like immunoreactivity in the same regions. The similar reductions of two prosomatostatin-derived peptides in AD cerebral cortex supports the contention that decreased somatostatin immunoreactivity in AD is caused by a degeneration of somatostatin cortical neurons and terminals.

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.

The products generated after addition of the ARG-LYS esteropeptidase activity purified from rat brain to synthetic somatostatin-28 were analyzed using radioimmunoassay, HPLC and amino acid analysis. In addition to somatostatin-14, both free arginine and free Lysine were identified together with somatostatin-28. The dipeptide ARG-LYS was not present, which indicates that three peptide bonds were hydrolyzed in order to achieve excision of the doublet. Since it is likely that the octacosapeptide is a precursor for both somatostatin-14 and somatostatin-28, these observations add further support to the hypothesis that the convertase is also involved in the in vivo processing of endogenous somatostatin-28.

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. Immunohistochemical visualization of hypothalamic neuronal cell bodies and fibers following cysteamine administration shows a disappearance of the SS14 immunoreactive fibers and cell bodies with no apparent change in the SS28(1–12) immunoreactive fibers and cell bodies. These data suggest that, in rat hypothalamus, selective release of SS14 and SS28(1–12) can occur. The results are discussed in relation to possible sites of storage and release of the somatostatin- related peptides from synaptic nerve terminals.

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. We could not detect SS14 immunohistochemically.

The primary structure of the NH2-terminally 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-Ala-Asn-Ser-Asn-Pro-Ala-Met-Ala-Pro-Arg-Glu-Arg-Lys-Ala-Gly-Cys-Lys-Asn-Phe-Phe-Trp-Lys-Thr-Phe-Thr-Ser-Cys-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.

Somatostatin-28 (SS28) is considered as a precursor of somatostatin-14 (SS14) and also of the remaining NH2 terminus of SS28, the dodecapeptide SS28 which has been recently characterized. In order to study the cellular and subcellular localization of SS28 in the rat hypothalamus, we have conducted a light and electron microscopic immunocytochemical study, using both the peroxidase-antiperoxidase and the immunogold technique. Several antisera which selectively recognize one or more of these 3 somatostatin-related peptides were used. In serial paraffin sections, it was observed that SS28 was contained in the same neuronal cell bodies which also contain SS28. These neurons were exclusively located in the periventricular nucleus. All the antisera used also produced a strong staining in the external zone of the median eminence. At the electron microscopic level, the 3 peptides were exclusively localized in all the dense core vesicles in cell bodies and axons in the periventricular nucleus and terminals in the median eminence. These results strongly suggest that SS28 and SS14 originate from the precursor SS28 and that processing of the precursor occurs in the cell body. They also support the hypothesis that the 3 peptides are released simultaneously under appropriate stimulation.

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.

We tested whether the vulnerability of somatostatin (SST) neurons in senile dementia of the Alzheimer type (SDAT) depended upon their co-localization with neuropeptide Y (NPY). Density estimates of SST28- and NPY-immunoreactive neurons and percentage of double-labeled SST-NPY neurons were obtained in the cortex (areas 9 and 25) and the bed nucleus of stria terminalis (BST), in 6 SDAT and 5 control cases. Counts of senile plaques (SP) and neurofibrillary tangles (NFT) were done on thioflavin S stains. In both cortical areas, a decrease in the density of SST28-IR neurons was found in SDAT cases (-60% in area 25 and -80% in area 9), whereas density of NPY-IR neurons was unchanged. Accordingly, the proportion of single-labeled SST neurons decreased; this decrease was significantly correlated with SP (r = -0.89, P less than 0.001). We conclude that single SST-IR neurons, in cortical layers II-III, and V, are preferentially lost relative to co-localized SST-NPY neurons. In the BST, no significant reduction of SST-IR, NPY-IR neurons nor of the percentage of single labeled SST neurons was found, despite the presence of SP. Thus one subpopulation of SST neurons, defined by associated neurochemical characters (not co-localized with NPY nor with NADPH diaphorase) and by topography (cortical layers III and V) appears to be particularly vulnerable in SDAT. The potential importance of their position in neural circuitry is emphasized.

The possible role of medullary somatostatin as a neurotransmitter involved in blood pressure regulation was examined. Blood pressure and heart rate were measured in pentobarbital anesthetized rats while pmol quantities of somatostatin-28 (SS28), SS28-(15–28) (somatostatin-14), SS28-(1–12) or SS28-1(1–10) were injected into the caudal nucleus tractus solitarii (NTS). Injection of artificial cerebrospinal fluid (ACSF; 500 nl/10 s) alone was followed by a small (2 mmHg) statistically insignificant decrease in blood pressure. SS28, SS28-(15–28) or SS28-(1–12) induced an immediate and statistically significant decrease in blood pressure as compared to vehicle alone. SS28-(1–10) had no significant effect. SS28 was tested at three doses. The lowest dose of SS28 (2.5 pmol) induced an 8 mmHg drop in blood pressure, the medium dose (25 pmol), an 11 mmHg drop and the highest dose (250 pmol) a 16 mmHg drop. The other peptide fragments were tested only at the higher 250 pmol dose. SS28-(1–12) induced a significantly (P < 0.01) less drop in blood pressure (10 mmHg) when compared to either SS28 or SS28-(15–28) (18–19 mmHg). The hypotensive and bradycardiac effects of SS28 were prevented by pretreatment with methylscopolamine (5 mg/kg i.p.) 10 min prior to SS28. The cardiovascular effects of intramedullary injected SS28 were also blocked with an ipsilateral NTS injection of yohimbine or tolazoline 10 min prior to SS28. These data indicate that the hypotension and bradycardia following the intramedullary injection of SS28 is mediated through the parasympathetic nervous system and that the C-terminal portion of SS28 and SS28-(15–28) is important for potency, but not obligatory, for efficacy. In addition, because two different adrenergic blocking agents can prevent the cardiovascular effects of somatostatin, central adrenergic mechanisms may be involved.

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.

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 somato-statin-28(1–2) like immunoreactivity (S-28(1–2) 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. Immuno-reactivity in the eluting fractions was assayed with two separate, region specific RIAs using antibodies R149 (S-14 LI) and S309 (S-28(1–2)LI). RIA R149 recognizes the 6-8 and 14 regions of the S-14 sequence and detects S-14, S-28, and prosomatostatin, an approximately 14,000 mol wt precursor for the two peptides. RIA S309 recognizes the 2–11 segment of S-28 and reacts with S-28, S-28(1–2), and higher mol wt S-28(1–2) LI but not S-14. Total tumor S-14 LI was 190 pmol/mg protein and consisted of three peaks of immunoreactivity of apparent 14,000 mol wt (14K S-14 LI), 3,200 mol wt (3.2K corresponding to...

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.

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.

Following the discovery of the growth hormone release-inhibiting factor somatostatin from extracts of ovine hypothalamus1, an N-terminally extended somatostatin of 28 amino acids has been identified in mammalian tissue2–6. The original peptide, somatostatin-14 (SS14), corresponds to the C-terminus of somatostatin-28 (SS28). Both SS28 and SS14 have biological activity3,7–9, occur in several rat brain regions10–15, are present in cell bodies and nerve terminals11–14,16 and can be released in vitro upon depolarization in a calcium-dependent manner16–19. Further, high-affinity binding sites were described for SS14, which also bind SS28 (refs 20–23). Recently, a dodecapeptide which corresponds to the N-terminus of somatostatin-28, somatostatin-28(1–12), has been characterized in rat hypothalamus24. Radioimmunological25 and immunohistochemical13 studies have indicated the presence of SS28(1–12)-like immunoreactivity in several cortical and subcortical regions of the rat brain. This peptide was found to be unevenly distributed with the highest concentration in the hypothalamus, and preferentially localized to dendritic and axonal processes and terminals. These observations suggest that SS28(1–12) may be a neurotransmitter. In this study, we describe a calcium-dependent release of a SS28(1–12)-like peptide from hypothalamic slices in vitro. This finding supports a neurotransmitter function for this peptide.

A peptide representing the major form of somatostatin-like immunoreactivity was isolated from 600 rat pancreata by using anti-somatostatin affinity chromatography, gel permeation chromatography and reverse-phase high-performance liquid chromatography (HPLC). The isolated peptide elutes with the same retention time as synthetic somatostatin-14 in isocratic HPLC and its amino acid composition is in agreement with that of the tetradecapeptide. We propose that the structure of the major rat pancreatic somatostatin is identical to that of somatostatin-14 characterized in other species.

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. In vitro cysteamine significantly increased the basal release of SS14-LI and markedly potentiated the evoked release of SS14-LI from hypothalamic slices. At 10 mM cysteamine, enhanced release of SS14-LI from hypothalamic slices was still observed despite a marked depletion of tissue content of SS14-LI.

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 125 I -N- Tyr-SRIF -14 and 125 I -N- Tyr-SRIF -28 are present on pituitary and brain membranes. Under the same conditions, 125 I -N- Tyr-SRIF -28(1–12) binding is undetectable. Moreover, SRIF-28(1–12) does not displace 125 I -N- Tyr-SRIF -14 or 125 I -N- Tyr-SRIF -28 binding. SRIF-28 is more potent than SRIF-14 to displace 125 I -N- Tyr-SRIF -28 binding to brain and pituitary membranes, while both peptides are equipotent to displace 125 I -N- Tyr-SRIF -14 binding. Finally, the regional distribution of 125 I -N- Tyr-SRIF -14 and 125 I -N- Tyr-SRIF -28 binding sites in the brain is identical. In conclusion, the present results are consistent with a neurotransmitter and neurohormonal role for SRIF-14 and SRIF-28. The function of SRIF-28(1–12) in brain remains to be elucidated. Additionally, a differential role for SRIF-14 and SRIF-28 both in adenohypophysis and brain cannot be ascertained at the present time.

Somatostatin-28 1 12 concentrations were measured in Huntington's disease (HD) postmortem tissue using a specific radioimmunoassay. Concentrations of immunoreactive somatostatin-28 1 12 were significantly increased in the caudate and putamen but were unchanged in cortical areas A9 and A17. Since somatostatin-28 1 12 terminates with the amino acids Arg-Glu-OH, we examined whether this dodecapeptide compound might exert a neurotoxic effect. Injections of somatostatin-28 1 12 into rat striatum showed no evidence of histologic damage.

Somatostatin is a tetradecapeptide known for its inhibitory action on the release of several hormones in addition to somatotropin (1,2).

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. The amino acid compositions of both peptides in pancreas and hypothalamus correspond to the fragment 1-12 of SS28. The more hydrophobic peptide from each tissue coeluted with synthetic SS28-(1-12) on HPLC, while the other one coeluted with synthetic SS28-(1-12)-amide. We conclude that the prosomatostatin fragment Ser-Ala-Asn-Ser-Asn-Pro-Ala-Met-Ala-Pro-Arg-Glu-OH is present in both rat hypothalamus and rat pancreas.

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. These observations show that the relative amounts of immunoreactive sites related to SS28 and its cleavage product SS28-(1-12) in maturing and mature secretory granules are different in pancreatic, gastric, and intestinal D cells.

Somatostatin (SOM) immunohistochemistry was used to map SOM-containing fibers, perikarya and terminal-like structures in the human forebrain of 8 control and 8 Alzheimer's diseased patients. Immunohistochemically processed tissue revealed a somatostatin-immunoreactive amygdalostriatal fiber pathway apparently originating from SOM-positive neurons of the central amygdaloid nucleus. This pathway coursed dorsomedially between the optic tract and the internal segment of the globus pallidus within the ansa peduncularis-ventral amygdalofugal fiber system en route to the substantia innominata-nucleus basalis complex, the bed nucleus of the stria terminalis, the medial preoptic hypothalamic area, the nucleus accumbens and the olfactory tubercle. Somatostatin fibers associated with this pathway appeared as coarse heavily stained 'wooly fibers'. At the light microscopic level, there was no apparent difference in this somatostatin containing amygdalostriatal pathway between neurologically normal and Alzheimer's diseased brains.

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. These data suggest that there may be an intraneuronal transformation from a SS28-like peptide to a SS28 (1-12)-like peptide. In addition, the enriched distribution of nerve fibers containing the antigenic determinant, SS28 (1-12), strongly implies that somatostatin-related peptides constitute a major neurotransmitter system in neocortex. The morphological characteristics of this system are homologous with long projection pathways such as the cortico-cortical specific intrinsic systems as well as projections.

Acid extracts of murine hypothalamic and extra-hypothalamic rat brains were analyzed by specific radioimmunoassays for the presence of somatostatin-14, somatostatin-28 and somatostatin-28(1–12)-like immunoreactivity. Seven molecular forms were observed after gel permeation chromatography. In addition to somatostatin-14, somatostatin-28 and somatostatin-28(1–12), there were two peptides of 4,400 and 7,500 mol. wt. which contained the somatostatin-28(1–12) sequence with an extension towards the NH2-terminus of pro-somatostatin. Moreover, two other peptides of 6,000 and 9,500 mol. wt. were detected containing the whole somatostatin-28 structure. These results imply that the processing of brain pro-somatostatin involves a minimum of four cleavage sites and yields at least seven peptides.

Two neuropeptides reacting with antibodies directed against the C-terminal region of somatostatin 281–12 were purified to homogeneity from rat brain extracts. Amino acid analysis revealed that the larger peptide (8 kdaltons) consisted of 76 amino acids. Microsequencing established its aminoterminal structure as: Ala-Pro-Ser-Asp-Pro-Arg-Leu-Arg-Gln-Phe-X-Gln-Lys. The 8 kdalton somatostatin 281–12-like peptide corresponds to the whole prosomatostatin molecule without Arg-Lys-somatostatin-14. The smaller peptide (5 kdaltons) consists of 44 amino acids and is generated after cleavage of a Leu-Leu bond at position 56–57 of pre-prosomatostatin. Both 8 kdalton and 5 kdalton somatostatin 281–12-like peptides contain somatostatin 281–12 at their C-termini. The 4 most abundant neuropeptides derived from pre-prosomatostatin (pre-proSS) and presently characterized are: somatostatin-14, somatostatin 281–12, somatostatin 28 and pre-proSS25–100.

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.