Somatostatin mRNA Expression Research Articles

Rapid reduction in somatostatin mRNA expression by hypothalamic neurons induced by dexamethasone.

We have previously reported that peripherally administered dexamethasone induces a rapid increase in hypothalamic somatostatin release. Here we investigated whether somatostatin synthesis could also be affected by this treatment and the potential involvement of glutamate in this effect. Male rats received a saline or a dexamethasone injection (300 microg/100 g body weight) and were killed 30 min later. Thirty minutes prior to dexamethasone treatment, another group received an i.p. injection of MK-801, a NMDA receptor antagonist. Cells expressing somatostatin mRNA in the periventricular nucleus were analyzed by in situ hybridization using digoxigenin-labeled somatostatin oligonucleotide probe. Dexamethasone decreased the number of digoxigenin-labeled cells expressing somatostatin mRNA in the periventricular nucleus as compared to the same histological sections from control rats. The dexamethasone effect was reversed by pretreatment with MK-801, which alone also decreased the number of cells expressing somatostatin mRNA. In summary, dexamethasone administration induces a significant rapid decrease in periventricular cells expressing somatostatin mRNA and this effect is partly abolished by MK-801.

Galanin-Like Peptide (GALP) mRNA Expression Is Restricted to Arcuate Nucleus of Hypothalamus in Adult Male Rat Brain

Galanin-like peptide (GALP) is a novel 60-amino acid neuropeptide, isolated from porcine hypothalamus and subsequently identified in rats and humans, which has reported selectivity for the Gal-R2 galanin receptor [Ohtaki T et al: J Biol Chem 1999; 274: 37041–37045]. In the current study, the regional and cellular distribution of GALP mRNA in rat brain has been investigated by in situ hybridization of [³⁵S]-labelled oligonucleotide probes. In a thorough screening of adult male rat brain, GALP mRNA expression was detected only throughout the rostrocaudal extent of the arcuate nucleus (ARC) with the most abundant hybridization signal in the posterior, periventricular zones. GALP mRNA-positive neurons were mostly localized in the ventromedial division of the ARC, with many closely adjacent to the wall of the third ventricle. Smaller numbers of labelled neurons were also found in ventrolateral areas. The distribution of GALP mRNA was somewhat complementary to that for galanin (GAL) mRNA in the ARC, but contrasted with the broad distribution of this transcript throughout the hypothalamus. GAL mRNA was also distributed along the rostrocaudal extent of the ARC, but was most abundant in the anterior to middle levels and in ventrolateral regions. Interestingly, somatostatin mRNA expression appeared to overlap the distribution of GALP mRNA in posterior, ventromedial regions of the ARC. Thus, in adult rat brain GALP mRNA expression was restricted to a discrete subpopulation of neurons in the ARC, with a unique localization pattern unlike GAL or many other known peptide- and transmitter-containing cells in this region. GALP could, however, be co-expressed in sub-populations of other neuronal phenotypes (e.g. somatostatin neurons) or within cells that express Gal-R2 receptors. In view of the established anatomy and function of the ARC and the restricted localization of GALP mRNA, this novel peptide is likely to play a role in regulation of anterior pituitary hormone secretion, or in regulation of other hypothalamic peptide and transmitter systems.

Anatomically specific changes in the expression of somatostatin, growth hormone-releasing hormone and growth hormone receptor mRNA in diabetic rats.

Growth hormone (GH) secretion is altered in poorly controlled diabetic animals. However, modifications in the hypothalamic neuropeptides that control GH secretion, somatostatin and GH-releasing hormone (GHRH), as well as changes in the sensitivity of the hypothalamus and pituitary to the feedback effects of GH, are less clear. We have used RNase protection assays and in-situ hybridization to address whether the mRNA expression of GH, somatostatin and GHRH, as well as of the GH receptor (GHR) in the hypothalamus and anterior pituitary, are altered in streptozotocin-induced diabetic rats. After induction of diabetes, rats were treated with insulin twice daily for 3 weeks to obtain either poorly controlled (mean plasma glucose >300 mg/dl) or well-controlled diabetic rats. Although no significant change in pituitary GH mRNA expression was found, the hypothalamic expression of GHRH and somatostatin mRNA was reduced in poorly-controlled diabetic rats and returned to control values with normalisation of plasma glucose concentrations (P<0.0001 and P<0.002, respectively). Somatostatin mRNA expression was reduced only in the central portion of the periventricular nucleus, with no change being seen in the other areas of the periventricular nucleus or in the arcuate, suprachiasmatic or paraventricular nuclei. A significant decline in GHRH mRNA expression was observed in both the arcuate nucleus and ventromedial hypothalamus. Anterior pituitary GHR mRNA expression was significantly reduced in both well and poorly-controlled diabetic rats, while there was no change in the hypothalamus. To examine whether the evolution time of the diabetes influences these parameters, in a subsequent experiment, diabetic rats received no insulin for 2 months. A significant decline in GHRH and somatostatin mRNA expression was also observed in these rats. In addition, pituitary GH mRNA expression declined significantly in long-term diabetic rats. These results demonstrate that: (1) the expression of both GHRH and somatostatin declines specifically in anatomical areas involved in anterior pituitary hormone control; (2) GHR mRNA expression is decreased in the pituitary of diabetic rats, but not in the hypothalamus, and does not return to control values with normalisation of mean blood glucose concentrations; and (3) the evolution time of the diabetes is important for detecting some changes, including the decrease in pituitary GH mRNA expression.

Acid-sensitive and alkaline-sensitive sensory neurons regulate pH dependent gastrin secretion in rat.

We examined the role of capsaicin-sensitive afferent neurons in pH-dependent gastrin secretion in the rat stomach. The change in serum gastrin levels relative to changes in luminal pH (using omeprazole for luminal alkalization or 0.1 N HCl for luminal acidification) was studied after oral administration of 4% lidocaine or capsaicin-induced ablation of afferent neurons. The increase of serum gastrin levels by luminal alkalization was significantly inhibited (50%) after administration of 4% lidocaine. Capsaicin pretreatment (125 mg/kg subcutaneously over two days) inhibited the change in serum gastrin levels both the luminal alkalization (38%) and acidification (66%). Antral gastrin contents, somatostatin contents, gastrin mRNA expression, and somatostatin mRNA expression were not significantly affected by capsaicin pretreatment. Our results indicate that capsaicin-sensitive afferent neurons participate in the secretion of gastrin by luminal alkalization and inhibition of gastrin by luminal acidification.

Spontaneous ECLomas in cotton rats (Sigmodon hispidus): tumours occurring in hypoacidic/hypergastrinaemic animals with normal parietal cells.

We have identified cotton rats with a high female-predominant occurrence of spontaneous gastric carcinomas localized to the oxyntic mucosa, classified as malignant enterochromaffin-like (ECL) omas. The present study was made to further characterize these ECLomas and surrounding oxyntic mucosa, both morphologically using histochemical and immunohistochemical methods, and for gene expression by northern blot analysis. Among eight female cotton rats, three had an irregularly thickened oxyntic mucosa, increased stomach weight and a high serum gastrin level. Histopathological examination showed adenomatous hyperplasia of the thickened oxyntic mucosa with areas of an invasive neoplastic tumour. Immunohistochemistry, using the general neuroendocrine cell marker chromogranin A (CgA) and the specific ECL cell marker histidine decarboxylase (HDC), showed a considerably increased ECL cell density. These ECL cells displayed active proliferation, with hyperplasia, dysplasia and neoplasia. Parietal cells were not found in the tumour tissue. Parietal cell density was only slightly reduced in the surrounding oxyntic mucosa. The antral mucosa was histopathologically normal with a normal number of gastrin-immunoreactive cells. Likewise, somatostatin-immunoreactive cells did not show any differences in the antral and oxyntic mucosa between rats with pathological and normal oxyntic mucosa. Northern blot analysis revealed increased expression of CgA and HDC mRNA in the thickened oxyntic mucosa, whereas H(+)/K(+) ATPase mRNA was similar in the oxyntic mucosa of those with thickened and normal oxyntic mucosa. Gastrin mRNA in the antral mucosa was high in animals with thickened oxyntic mucosa. Somatostatin mRNA expression was similar in the antral mucosa of control animals and animals with a thickened oxyntic mucosa. We conclude that the spontaneous gastric carcinoma occurring in female cotton rats is an ECLoma developing secondary to hypergastrinaemia due to reduced intragastric pH. The mechanism for reduced acidity is not known, but is not gastric atrophy.

Correlation of hypothalamic somatostatin mRNA expression and peptide content with secretion: sexual dimorphism and differential regulation by gonadal factors.

Sex differences in growth hormone (GH) secretion in the rat are thought to be determined, to a large extent, by gonadal steroid-dependent sex differences in somatostatin (SRIH) secretion from neurones in the periventricular nucleus (PeN) which project to the median eminence (ME). The present study aimed to obtain direct evidence for sex differences and gonadal regulation of SRIH release within this pathway and to determine the relationships between SRIH mRNA expression, SRIH peptide content and SRIH secretion in the adult rat. Somatostatin mRNA expression in the PeN and peptide content in both PeN and ME were higher in males than females (P<0.05). However, both basal and 56 mM K+-stimulated SRIH release in vitro from hypothalamic explants incorporating the PeN-ME pathway were higher (P<0.01) in females. The gonadectomy of female rats resulted in significantly reduced basal levels of SRIH release equivalent to that of males but had no effect on SRIH mRNA/peptide content or K+-stimulated release. In contrast, gonadectomy of male rats reduced SRIH mRNA and peptide contents and elevated K+-stimulated secretion (P<0.01) to levels similar to that seen in intact females, without affecting basal release. In summary, these results demonstrate that in the PeN-ME of the adult rat: (1) SRIH mRNA and peptide content is well correlated and sexually dimorphic but dependent on gonadal factors in the male only; (2) SRIH secretion is sexually dimorphic and dependent on gonadal factors; but (3) differences in mRNA/peptide content do not reflect secretory capacity; and (4) gonadal factors differentially modulate SRIH secretory dynamics in males and females.

Estrogen-dependent ontogeny of sex differences in somatostatin neurons of the hypothalamic periventricular nucleus.

The sexually dimorphic profile of GH secretion is thought to be engendered by gonadal steroids acting in part on hypothalamic periventricular somatostatin (SOM) neurons. The present study set out to examine and characterize the development of sex differences in these SOM neurons. In the first series of experiments, we used in situ hybridization to examine SOM messenger RNA (mRNA) expression within the periventricular nucleus (PeN) of male and female rats on postnatal day 1 (P1), P5, and P10. Cellular SOM mRNA content was found to increase from P1 to P10 in both sexes (P < 0.01), but was 24% (P < 0.05) and 38% (P < 0.01) higher in males on P5 and P10, respectively. A second series of experiments examined the SOM peptide content of the PeN in developing rats and found increasing levels from P1 to P10, with a 44% higher SOM content in males compared with females on P10 (P < 0.05). The third series of experiments questioned the role of gonadal steroids in engendering sex differences in SOM mRNA expression by determining the effects of neonatal gonadectomy (GDX) and replacement of dihydrotestosterone or estradiol benzoate. The SOM mRNA content of PeN neurons in P5 males gonadectomized on the day of birth was the same as that in P5 females and was significantly reduced compared with that in sham-operated P5 males (P < 0.05). Male rats GDX on P1 and treated with estradiol benzoate from P1 to P5 had cellular SOM mRNA levels similar to those in intact males on P5, whereas dihydrotestosterone treatment had no effect. Treatment of intact males with an androgen receptor antagonist, cyproterone acetate, on P1 had no effect on cellular SOM mRNA on P5, whereas male rats given the aromatase inhibitor 1,4,6-androstatriene-3,17-dione from P1 to P5 had lower (P < 0.05) SOM mRNA levels than controls. In the final set of experiments, dual labeling immunocytochemistry showed that SOM neurons in the PeN of P5 rats did not contain estrogen receptor-alpha, but expressed androgen receptors in a sexually dimorphic manner. These results demonstrate that a sex difference in SOM biosynthesis, which persists into adulthood, develops between P1 and P5 in PeN neurons. Despite the absence of estrogen receptor-alpha in these neurons, the organizational influence of testosterone only occurs after its aromatization to estrogen.

Expression of somatostatin mRNA in various differentiated types of gastric carcinoma.

AIM:To investigate the expression of somatostatin mRNA in various differentiated types of gastric carcinoma.METHODS: By using in situ hybridization and immunohistochemical techniques, the expression of somatostatin mRNA and somatostatin immunoreactivity in the normal gastric mucosa, the poorly, moderately and well-differentiated gastric carcinomas, and various clinical stages of carcinoma were observed.RESULTS: In comparison with the normal gastric mucosa, the significantly increased expression of somatostatin mRNA positive cells was displayed in gastric carcinoma (t = 2.681, P < 0.01). The positive signal cells were distributed in a scattered form or aggregated as a mass or a cord, and the positive cells were more significantly enhanced in poorly differentiated carcinomas than those in well and moderately differentiated carcinomas (t = 2.962, P < 0.01). The somatostatin mRNA hybridization signals in stages III and IV of gastric carcinoma were significantly higher than those in stages I and II. The results of somatostatin immunoreactivity were consistent with those of in situhybridization.CONCLUSION: The alteration of the expression of somatostatin mRNA was associated with the development of gastric carcinoma and may play an important role in theprocess of tumor differentiation.

Expression of antral somatostatin mRNA in duodenal ulcer disease complicated with antral gastritis associated withHelicobacter pylori

Expression of antral somatostatin mRNA in duodenal ulcer disease complicated with antral gastritis associated with Helicobacter pylori

Potentiating Hypergastrinemic Effect by the Peroxisome Proliferator Ciprofibrate and Omeprazole in the Rat

Background: Profound inhibition of gastric acid secretion induces enterochromaffin-like (ECL) cell carcinoids due to hypergastrinemia. Peroxisome proliferators also lead to hypergastrinemia and ECL cell carcinoids but without reducing gastric acidity. Since the peroxisome proliferator ciprofibrate is still in use as lipid-reducing agent, and proton pump inhibitors are among the most commonly used drugs, we found it of interest to evaluate both the effect of a combination of these drugs on serum gastrin and the expression of gastrin and somatostatin mRNA in antral mucosa. Methods: The drugs were given by gastric gavage once daily for 4 weeks to female rats. Blood was drawn by vein puncture before and at the end of the 4-week period for determination of gastrin by radioimmunoassay. At death the stomachs were removed, the antral mucosa homogenized, and the density of gastrin and somatostatin mRNA determined by Northern blot, using 32P-labelled probes. Results: Omeprazole dosing increased serum gastrin 4-fold, ciprofibrate 5-fold, and the combination 24-fold. Serum gastrin during ciprofibrate dosing increased gradually, reaching significance after 14 days. Antral gastrin mRNA density increased similarly to the increase in serum gastrin, whereas antral somatostatin mRNA tended to be reduced in the omeprazole and increased in the ciprofibrate-dosed rats. Conclusion: A potentiating hypergastrinemic effect of the peroxisome proliferator ciprofibrate and the inhibitor of gastric acid secretion omeprazole is shown, indicating different mechanisms of action.

Fatty Acids Decrease IDX-1 Expression in Rat Pancreatic Islets and Reduce GLUT2, Glucokinase, Insulin, and Somatostatin Levels

IDX-1 (islet/duodenum homeobox-1) is a transcription factor expressed in the duodenum and pancreatic beta and delta cells. It is required for embryonic development of the pancreas and transactivates the Glut2, glucokinase, insulin, and somatostatin genes. Here we show that exposure of isolated rat pancreatic islets to palmitic acid induced a approximately 70% decrease in IDX-1 mRNA and protein expression as well as 40 and 65% decreases in the binding activity of IDX-1 for its cognate cis-regulatory elements of the Glut2 and insulin promoters, respectively. The inhibitory effect of palmitic acid required its mitochondrial oxidation since it was prevented by the carnitine palmitoyltransferase I inhibitor bromopalmitic acid. The palmitic acid effect on IDX-1 was correlated with decreases in GLUT2 and glucokinase expression of 40 and 25%, respectively, at both the mRNA and protein levels. Insulin and somatostatin mRNA expression was also decreased by 40 and 60%, whereas glucagon mRNA expression was not modified. After 48 h of exposure to fatty acids, total islet insulin, somatostatin, and glucagon contents were decreased by 85, 55, and 65%, respectively. At the same time, total hormone release was strongly stimulated (13-fold) for glucagon, whereas its was only marginally increased for insulin and somatostatin (1.5- and 1.7-fold, respectively). These results indicate that elevated fatty acid levels 1) negatively regulate Idx-1 expression; 2) decrease the expression of genes transactivated by IDX-1 such as those for GLUT2, glucokinase, insulin, and somatostatin; and 3) lead to an important increase in glucagon synthesis and secretion. Fatty acids thus have pleiotropic effects on pancreatic islet gene expression, and the negative control of Idx-1 expression may be an initial event in the development of these multiple defects.

Selective increase in somatostatin mRNA expression in human basal ganglia in Parkinson's disease

Levels of the neurotransmitter somatostatin (SS) have previously been shown to be reduced in the cortex and hippocampus of demented parkinsonian patients and patients with Alzheimer's disease. In situ hybridisation histochemistry (ISHH) was performed with an 35S tail-labelled oligonucleotide DNA probe to human SS mRNA, to examine its expression within the striatum, medial medullary lamina (MML) and reticular thalamic nucleus in Parkinson's disease (PD) and in matched controls. A chronic unilaterally MPTP-lesioned L-DOPA-naive primate model was also examined for comparison of SS mRNA expression with that in human L-DOPA treated PD subjects. Quantitation of SS mRNA expression on emulsion dipped sections revealed a significant increase (82%) in the MML of the globus pallidus in PD (56.5 μm 2 of silver grain/cell, n=9 cases) compared to controls (26.3 μm 2/cell, n=13 cases, p<0.01, Student's t-test), paralleling the increase previously observed by this group for NOS mRNA. SS mRNA expression was higher in the dorsolateral than ventromedial putamen in controls ( p<0.001; DL: 24.89±SEM 1.35; VM: 17.96±SEM 2.63; n=14) but this gradient was lost in PD cases ( p>0.05; DL: 22.68 ± 1.94; VM: 22.17 ± 2.94; n=10). These findings suggest specific modification of basal ganglia SS-ergic pathways in PD.

Ischemia changes the coexpression of somatostatin and neuropeptide Y in hippocampal interneurons.

Transient cerebral ischemia causes extensive cell death in hippocampal CA1 pyramidal cells and selective loss of interneurons in the dentate hilus. Many hippocampal interneurons can be classified by their contents of somatostatin (SS) and/or neuropeptide Y (NPY). Following ischemia in the rat, most of the NPY immunoreactivity is permanently lost in hippocampus. Furthermore, SS interneurons in the dentate hilus die, whereas CA1 interneurons survive and their expression of SS mRNA and peptide returns to preischemic levels within 16 days after ischemia. We have addressed the following questions: (1) Does the loss of NPY involve a specific downregulation in surviving CA1 interneurons that pre-ischemically expressed both SS and NPY? (2) Can the subpopulation of dying interneurons in hilus be identified from their preischemic coexpression of SS and NPY? We investigated the coexpression of SS mRNA and NPY peptide using combined in situ hybridization and immunocytochemistry. Cells containing one or both markers were counted in control sections and sections taken 2-16 days after ischemia from the hippocampal formation. In CA1, a decrease in the number of neurons containing NPY alone as well as a decrease in the number of neurons coexpressing NPY and SS was observed, whereas the number of neurons containing SS alone increased 16 days after ischemia. We conclude that neurons coexpressing SS and NPY before ischemia added to the number of neurons containing SS alone after ischemia, because NPY expression was selectively down regulated in the coexpressing population. In hilus, we demonstrated both survival and ischemic cell death of neurons expressing either SS, NPY or both, indicating that hilar interneurons dying from ischemia cannot unequivocally be identified from their preischemic colocalization of SS and NPY.

Effects of Fasting on the Expression of Gastrin, Cholecystokinin, and Somatostatin Genes and of Various Housekeeping Genes in the Pancreas and Upper Digestive Tract of Rats

It is generally accepted that while the gene expression of many gut hormones in the pancreas and upper digestive tract is influenced by the prandial state, the expression of house-keeping genes (used as internal standards) is stable. We have analysed how food deprivation affects the messenger (m) RNA expression of gastrin, cholecystokinin (CCK), and somatostatin and of house-keeping genes glyceraldehyde-3-phosphate dehydrogenase (GAPDH), β-actin and 18S ribosomal (r) RNA. RNA, isolated from oxyntic, antral and duodenal mucosa and pancreas, was subjected to Northern blot analysis using complementary RNA probes. Compared to fed rats, food-deprived rats exhibited reduced mRNA expression of gastrin (antrum), somatostatin (antrum), CCK (duodenum), GAPDH (all tissues studied) and β-actin (all tissues studied) and unchanged 18S rRNA expression. We conclude that the assessment of gut hormone mRNA expression may be greatly influenced by the choice of internal standard and that 18S rRNA is superior.

Identification of growth hormone-releasing hormone and somatostatin neurons projecting to the median eminence in normal and growth hormone-deficient Ames dwarf mice.

In the spontaneous mutant Ames dwarf mouse, GH deficiency coincides with a dramatic increase in the expression of both mRNA and peptide for stimulatory GHRH and reduced expression of GH-inhibitory somatostatin (SRIH) mRNA and peptide. However, both GHRH and SRIH are markedly reduced in the dwarf median eminence (ME), suggesting that ME innervation by GHRH and SRIH neurons may be aberrant in the absence of GH. In order to test this hypothesis, the number of GHRH and SRIH ME-projecting neurons was evaluated in normal and dwarf mice using a combination of retrograde tract-tracing and neuron phenotype identification by immunocytochemistry (ICC). Adult animals were injected intraperitoneally with the fluorescent tract-tracer fluorogold (FG), which, in the brain, is taken up only by axons terminating in areas deprived of the blood-brain barrier, such as the ME. Visualization of FG was achieved by either UV illumination or ICC, and was combined as appropriate with fluorescence or bright-field ICC for GHRH or SRIH. Cells immunoreactive for GHRH or SRIH and labeled with FG were quantified at each 180-microns rostral-to-caudal level through the hypothalamus. As reported previously, the total number of hypophysiotropic GHRH neurons was markedly increased in dwarf compared with that in normal mice. However, a similar percentage of ME-innervating GHRH cells was estimated in dwarf (73 +/- 4%) and normal (76 +/- 3%) animals. Such a percentage in dwarfs thus represents a larger population of ME-projecting GHRH cells (749 +/- 53) than in normal mice (128 +/- 15). Increased numbers of FG-labeled GHRH neurons in dwarfs were located at the middle and posterior levels of the arcuate nucleus (2.08, 2.26 and 2.44 mm posterior to bregma). The percentage of FG-labeled SRIH neurons was also similar for dwarf (83 +/- 2%) and normal (87 +/- 2%) mice. Because the total SRIH-immunoreactive neuronal population in dwarfs is significantly reduced compared to that in normal animals, the similar FG-labeled percentage reflects a reduced number of SRIH cells projecting to ME in dwarf (1,376 +/- 104) compared with normal (3,192 +/- 267) mice. Fewer FG-labeled SRIH cells were found in dwarfs at every anterior-to-posterior level of the periventricular nucleus (p < 0.01 for comparisons at 0.28, 0.46, 0.64, and 1.0, and p < 0.05 for comparison at 1.18 mm posterior to the bregma). The present study indicates that the reduction in GHRH and SRIH immunoreactivity in the dwarf ME may result from different phenomena for each neuronal population. The reduction in GHRH immunostaining in the ME, despite a marked increase in the total ME-projecting GHRH neurons, may be interpreted as increased GHRH release, with consequent depletion of the ME stores. In contrast, the deficit in ME SRIH may be proportional to the deficit in the number of detectable SRIH periventricular nucleus neurons.

Presence of somatostatin sst 2 receptors in the developing rat auditory system

A transient expression of somatostatin mRNA as well as of the peptide itself has been described in the developing mammalian auditory brainstem. However, little is known about the presence, and the spatial and temporal pattern of somatostatin (SRIF) receptor subtypes in this system. Therefore, we investigated the distribution of SRIF receptor binding sites labeled with the radioligands [ 125I]LTT-SRIF-28, [ 125I]Tyr 3-octreotide, and [ 125I]CGP 23996 (in buffers containing either Mg 2+ or Na + ions) within the developing auditory brainstem of the rat. In addition, we performed in situ hybridization with a 33P-labeled oligoprobe, specific for somatostatin sst 2 receptor mRNA. We observed a transient expression of SRIF receptors, labeled with [ 125I]LTT-SRIF-28, [ 125I]Tyr 3-octreotide, and [ 125I]CGP 23996 (only in the presence of Mg 2+ ions), in all principal auditory nuclei during neonatal development. In the adult rats, however, only the inferior colliculus displayed significant SRIF receptor binding. A very similar spatiotemporal labeling pattern was found for sst 2 receptor mRNA. Our in situ hybridization data, together with those on ligand binding, suggest a predominantly transient expression of sst 2 receptors in the auditory system. Since sst 2 sites (and possibly sst 3 and sst 5) as well as SRIF itself appear to be co-expressed during a period when synapse maturation occurs, we suggest that sst 2 receptors are involved in this process of the developing auditory system.

In situ hybridization for somatostatin mRNA in the adult rat: cingulate, insular, prepiriform, perirhinal, entorhinal, and retrosplenial cortical regions.

The expression of somatostatin mRNA within the allocortex of the rat was examined by in situ hybridization with an alkaline phosphatase labeled probe. We sought to determine whether parcellation of the allocortex could be based upon the number and laminar location of the hybridized cells and to contrast the allocortical features with those of the isocortical areas. The cingulate region was characterized by intense, moderate, and faint cells, small to medium in size throughout the laminae. The retrosplenial region demonstrated a somewhat stratified appearance with an abundance of cells expressing somatostatin mRNA in the upper portion of the composite layer II-IV and also in the upper portion of layer VI. The insular region displayed more heterogeneity. The distribution of the cells hybridized for somatostatin mRNA formed distinctive configurations within the insular region (dorsal and ventral agranular insular areas) with no obvious generality. The perirhinal area resembled the ventral agranular insular area, and the cell distribution of the entorhinal and prepiriform areas displayed a common characteristic in that the primary axis of the perikarya of somatostatin mRNA expressing cells within the lower layers were oriented at almost every possible angle. The conclusion of the investigation is that in situ hybridization for somatostatin mRNA provides a means by which the areal boundaries within the allocortex may be drawn.

Growth hormone-releasing factor expression is discordantly regulated in the hypothalamus and testis of streptozotocin-diabetic rats

Growth hormone-releasing factor (GRF) mRNA expression in male rats occurs predominantly in the hypothalamus (mainly in the arcuate nucleus), and among extraneural sites primarily in the testis. Hypothalamic GRF is the physiological tropic stimulus to growth hormone secretion. However, the role of GRF in the testis is unknown. We have shown previously that hypothalamic GRF mRNA expression is significantly reduced in streptozotocin (STZ)-diabetic rats. This reduction is confined to the arcuate nucleus and probably accounts for the suppression of growth hormone pulsatility. The present studies were performed to evaluate GRF expression in the testis of streptozotocin (STZ)-diabetic rats. Diabetes was induced by injection of STZ (100 mg/kg i.p.). Seventeen to twenty days later diabetic rats were hyperglycemic compared with vehicle-injected controls and demonstrated growth failure. Insulin treatment reduced the glycemia and increased body weight towards normal. Total RNA was extracted from the hypothalamus and testis, and GRF mRNA levels estimated by solution hybridization/nuclease protection assay. Levels of hypothalamic somatostatin mRNA were measured to serve as control values. GRF mRNA was significantly (P < 0.001) decreased in the hypothalamus of STZ-diabetic rats (0.2 +/- 0.07 mean relative densitometric units, n = 8) compared with controls (1.0 +/- 0.19, n = 8) with no change in somatostatin mRNA expression. In contrast, testicular GRF mRNA was increased 70% (P < 0.05) in STZ-diabetic rats. Insulin treatment resulted in normalization of hypothalamic GRF mRNA levels (1.1 +/- 0.17, n = 5) with no effect on testicular GRF mRNA expression. In conclusion GRF gene expression is discordantly regulated in tissues of male STZ-diabetic rats. While reduced GRF expression may account for the low growth hormone state in this model, increased testicular GRF mRNA (with the previously reported reduction of insulin-like growth factor-I mRNA) resembles the response seen in growth hormone-sensitive tissue (especially the hypothalamus) to this growth hormone-deficient state.

Embryonic and postnatal mRNA distribution of five somatostatin receptor subtypes in the rat brain

The messenger RNA (mRNA) expression of somatostatin (SRIF) receptors SSTR-1, SSTR-2, SSTR-3, SSTR-4 and SSTR-5 (called sst 1–5, now) was studied in rat brain between embryonic day 17 (E17) and post-natal day 5 (P5) by in situ hybridization histochemistry and compared to that of adult rats, sst 1 receptor mRNA expression was very low and restricted at E17, spread out at E18, to reach very high levels comparable to that of adult at P5 (e.g. in temia tecta, posteromedial cortical amygdaloid nucleus, subiculum). At E17 E18 , sst 2 receptor mRNA expression was low and limited (telencephalon); significant levels were present at P5 in allocortex, hippocampus, locus coeruleus, similarly to adult brain. sst 3 receptor mRNA was high at E17 in most brain regions, and almost as ubiquitous as in adult brain at P5. sst 4 receptor mRNA was apparently absent at E17, with low levelis in the hippocampus, amygdala and habenula at E18; a wider distribution, especially in the hippocampus and cerebral cortex was observed at P5, similar to that of adult, sst 5 receptor mRNA was not detected at E17 and negligible at E18; low levels were found in the cortex, hippocampus and cerebellum at P5. However, in adult brain, only the cerebellum and hind-brain showed some sst 5 receptor mRNA transcripts. The presence and distribution of SRIF receptor mRNAs differs substantially in embryo and adult brain. Some mRNAs are present throughout development, while others proceed only postnatally to the adult form. There are striking differences within and between the different SRIF receptor mRNAs, suggesting a role in neurogenesis for some SRIF receptors (e.g. sst 2). However, mRNA and protein levels do not necessarily correlate.

Somatostatin-mRNA expression in brainstem projections into the medial preoptic nucleus.

The medial preoptic nucleus plays an important role in the regulation of neuroendocrine processes, vegetative functions, sexual behaviour and the modulation of the somatomotoric system. The connections of the medial preoptic nucleus to other areas of the central nervous system are very complex, and the area receives afferents using numerous transmitters and neuropeptides. Previous investigations have shown that this nucleus receives afferents from various brainstem nuclei that also contain somatostatinergic neurons. This study was carried out to investigate if somatostatin-projecting neurons of the brainstem are afferents to the medial preoptic nucleus. This was approached by combining somatostatin-mRNA in situ hybridisation with True Blue retrograde tracing. Our results demonstrate somatostinergic brainstem projections into the medial preoptic nucleus mainly in the pedunculopontine nucleus and in the nucleus of the solitary tract (50% together). Other important somatostatinergic afferents into the medial preoptic nucleus originate in the cuneiform area, the dorsal parabrachial nucleus and in the lateral reticular nucleus (37% together). Less important are the somatostatinergic projections coming from the central grey, the laterodorsal tegmental nucleus, the locus coeruleus and the nucleus raphe magnus. Considering that these areas are involved in diverse functions such as cardiovascular regulation (nucleus of the solitary tract), transmission of visceral sensibility (dorsal parabrachial nucleus), modulation of the somatomotoric system (pedunculopontine nucleus) and in the regulation of neuroendocrine mechanisms (locus coeruleus), it seems tenable that the somatostatin projections demonstrated here also have a diverse functional quality within the medical preoptic nucleus where they terminate.