SRIF Neurons Research Articles

Immunolocalization of high molecular weight kininogen (HKg) and T kininogen (TKg) in the rat hypothalamus.

Specific HKg immunostaining detected with antiserum against the light chain (LC) of HKg was restricted to SRIF neurons of the hypothalamic periventricular area projecting to median eminence (ME). Heavy chain (HC) immunoreactivity related to HKg and/or low molecular weight kininogen (LKg) was found in some other hypothalamic territories. Specific TKg was mainly associated with vasopressin in neurons of suprachiasmatic (SCN), supraoptic (SON) and paraventricular (PVN) nuclei. By direct RIA, hypothalamus was found to contain the highest level of TKg (10ng/mg protein) and after trypsin hydrolysis and HPLC separation of kinins, 10.3 pg BK and 7.3 pg T-kinin/mg protein.

Brain somatostatin receptors in spontaneously hypertensive rats: An autoradiographic study

The apparent densities of brain somatostatin (SRIF) receptor sites were compared in adult spontaneously hypertensive rats (SH) and their normotensive genetic counterparts (Wistar-Kyoto; WKY) using quantitative receptor autoradiography. Globally, the distribution of brain [ 125I][Tyr 0,D-Trp 8]SRIF14 binding sites was very similar in both strains. However, apparent densities of specific labeling were either higher (subfornical organ, 3.2×; locus coeruleus, 1.9×; lateroanterior hypothalamic nucleus, 1.3×) or lower (basolateral amygdaloid nucleus, 0.8×; spinal trigeminal sensory nucleus, 0.6×) in SH than WKY rats in areas especially relevant to CNS cardiovascular integration. This provides further evidence for the possible involvement of brain SRIF neurons in cardiovascular regulation.

A Possible Role of Hypothalamic Somatostatin in the Maintenance of Rat Pituitary Responsiveness to Growth Hormone-Releasing Factor*

To characterize the role of hypothalamic somatostatin (SRIF) in regulating pituitary responsiveness to GH-releasing factor (GRF) in vitro, we reduced SRIF input to the rat anterior pituitary through the portal vessels. Three different paradigms were used as follows: 1) anterolateral hypothalamic deafferentation, 2) electrolytic lesions of the periventricular nucleus, and 3) passive immunization with SRIF antiserum. Rat CRF content in the stalk-median eminence markedly decreased to 19% and 57% of that of sham-operated controls 10 days after the deafferentation and the lesions, respectively. In contrast, rat GRF content was unchanged by either operation. SRIF content markedly decreased to 78%, 12%, and 2% of the control level 1, 3, and 10 days after deafferentation, respectively, and to 48% and 8%, 1 and 10 days after the lesions, respectively. The serum GH concentration was significantly increased 1 and 3 days after the deafferentation (P less than 0.01) and also 1 day after the lesions (P less than 0.01), followed by no increase 10 days after either operation. Anterior pituitary weight and GH content markedly decreased 3 and 10 days and 10 days after the deafferentation and the lesions, respectively. The human GRF (0.1 microM)-induced GH release response of anterior pituitaries removed from these treated rats was examined in an in vitro perifusion system. Even 1 day after these treatments, GH responsiveness was clearly attenuated by anterolateral hypothalamic differentiation (8.61 +/- 0.78 vs. 3.62 +/- 0.54 micrograms GH/h; P less than 0.01), periventricular nucleus lesions (6.52 +/- 1.07 vs. 3.20 +/- 0.53 micrograms GH/h; P less than 0.01) and passive immunization with SRIF antiserum (5.80 +/- 0.43 vs. 2.54 +/- 0.16 micrograms GH/h; P less than 0.01). This attenuated responsiveness gradually deteriorated 3 and 10 days after the surgical operations. These results indicate that SRIF neurons in the anterior periventricular nucleus play a role in maintaining the pituitary responsiveness to GRF, in addition to the original action of inhibiting GH release.

Characterization of the Glucose-Dependent Release of Growth Hormone-Releasing Factor and Somatostatin from Superfused Rat Hypothalami

The glucose-dependent secretion of the neuropeptides, growth hormone-releasing factor (GRF) and somatostatin (SRIF), by hypothalamic fragments was studied in vitro using a superfusion system. After equilibration of mediobasal hypothalami in HEPES-buffered Krebs-Ringer solution containing 5.5 mM glucose, glucose levels in the superfusion medium were altered. Lowering the glucose concentration in the medium from 5.5 to 2.7 or 1.1 mM provoked a rapid increase in GRF and SRIF release in a concentration and Ca2+-dependent manner. At 1.1 mM glucose, neuropeptide secretion was elevated 3- to 4-fold. The increase of GRF and SRIF release induced by low glucose was transient since stimulated neuropeptide secretion declined to basal levels in the continued presence of low glucose. Furthermore, after reequilibration in 5.5 mM glucose, no second stimulation of neuropeptide release could be induced by reduced glucose. Intracellular glucopenia induced by addition of 2-deoxy-D-glucose (16.5 mM) to the superfusion medium containing 5.5 mM glucose, also evoked increases in GRF and SRIF release. The sensitivity of GRF and SRIF neurons to glucose was absent in the postnatal period until day 9 after birth and then gradually increased. The parallel increases of GRF and SRIF release in response to low glucose observed in the present in vitro study, together with the suppression of plasma GH levels occurring in hypoglycemia in the rat, suggest that, in this condition, the inhibition of GH release induced by elevated SRIF levels predominates whereas the increase of GRF release might serve to attenuate this effect of SRIF.

NEURAL CIRCUITRIES RESPONSIBLE FOR THE REGULATION OF GROWTH HORMONE SECRETION IN THE RAT

Growth hormone (GH) secretion is mainly regulated by growth hormone releasing factor (GRF) and somatostatin (SRIF). Their chemical structures have already been determined and the distributions of immunoreactive GRF and somatostatin neurons in the central nervous system have been revealed. At this time, however, the neural mechanisms which regulate these two peptidergic neurons in the central nervous system are not fully understood. To clarify the neural circuitries responsible for the regulation of growth hormone secretion, electrical stimulation was delivered to the basolateral amygdala (ABL) and several brainstem nuclei in pentobarbital anesthetized rats.Electrical stimulation of the ABL caused an increase in the plasma GH level 10 min after the onset of stimulation. This increase in plasma GH level was not affected by prior lesioning of the hypothalamic periventricular nucleus (Pe), from which the major SRIF immunoreactive fibers in the median eminence originate. These results indicate that the increase in plasma GH level caused by electrical stimulation of the ABL is not caused by reducing SRIF secretion. This indicates that the ABL stimulation causes the excitation of GRF neurons and consequently induces GH secretion.Stimulation of the central gray matter in the midbrain and the pons, the locus ceruleus, the solitary tract nucleus and the lateral reticular nucleus suppressed exogenous GRF-induced GH secretion, but prior lesioning the Pe abolished the suppression of the GH secretion. These results indicate that electrical stimulation of several brainstem nuclei suppressed GH secretion through the excitation of SRIF neurons in the Pe.

Somatostatin-containing neuron systems in the rat hypothalamus: retrograde tracing and immunohistochemical studies.

By employing a combination of the immunohistochemistry for somatostatin (SRIF) and retrograde tracing with biotinylated wheat germ agglutinin (b-WGA) injected into the posterior pituitary (group 1) or into the median eminence (group 2), functional topography of hypothalamic SRIF neurons was determined in the rat hypothalamus. In group 1, large numbers of WGA-labeled neurons appeared in the rostral periventricular region and in the magnocellular division of the paraventricular and supraoptic nuclei; none of them were SRIF immunoreactive. In group 2, WGA-labeled neurons were numerous in the rostral periventricular region, the parvicellular division of the paraventricular nucleus, and the arcuate nucleus; most of the WGA-labeled neurons in the rostral periventricular region and some in the paraventricular nucleus were SRIF immunoreactive, but none in the arcuate nucleus showed immunoreactivity for SRIF. It is concluded that, in the rat hypothalamus, the locations of neurons containing hypophysiotrophic SRIF are confined within the rostral periventricular region and the parvicellular paraventricular nucleus. Our results do not support previous suggestions that SRIF immunoreactive axons innervate the posterior lobe of the pituitary.

Development of somatostatin immunoreactive neurons in the rat occipital cortex: a combined immunocytochemical-autoradiographic study.

The postnatal development of somatostatin (SRIF)-immunoreactive neurons, previously labeled with [3H]thymidine on embryonic days E14-E22, has been studied in the rat occipital cortex. Immunocytochemistry combined with autoradiography showed an "inside-out" maturation pattern. Only SRIF neurons generated at E14 were present in layer VI in newborn rats. Later generated SRIF neurons appeared progressively higher in the cortex until about postnatal day 12 when SRIF neurons from E21 appeared in layer II. At 2 weeks of age, therefore, all SRIF neurons from E14-E21 were present. Most of these had been generated between E15 and E17 with a moderate number at E14 and rapidly diminishing numbers from E18 to E21. Although an overall layered distribution was apparent at peak production, there was a tendency for diffuse distribution most noticeable at E17. Diffusely distributed neurons were more likely to be below their appropriate layer than above it, thus contributing extra SRIF neurons to layer VI. At 3, 4, and 5 weeks, progressively fewer SRIF neurons were seen with a consequent reduction in the number of double-labeled neurons. It is suggested that the transient population of SRIF neurons thus revealed plays a role in cortical development.

Immunohistochemical Identification of Somatostatin-Containing Neurons Projecting to the Median Eminence of the Rat

Immunohistochemical staining of somatostatin (SRIF) and a retrograde transport method with horseradish peroxidase (HRP) were simultaneously applied to the same section of the rat brain to identify the specific SRIF-containing neurons sending their fibers to the median eminence. After HRP injection into the median eminence, SRIF-positive neurons in the rostral parts of the periventricular nucleus were shown to be labeled with HRP, SRIF neurons in the other brain areas, such as the amygdala, the ventromedial nucleus, dorsomedial nucleus and arcuate nucleus, had no HRP-positive material. The present findings appear to demonstrate that SRIF neurons in the periventricular nucleus project directly to the median eminence.

Rostromedial septal area controls pulsatile growth hormone release in the golden hamster

Limbic forebrain inhibits growth and growth hormone (GH) secretion in mature golden hamsters as shown by acceleration of growth and increases in serum GH concentrations following the electrolytic lesions of septum, transection of the hippocampus and surgical separation of these two regions. The growth-inhibitory function of this circuit is most probably mediated by somatostatinergic (SRIF) neurons. Such lesions induce hypoactivity possibly due to damage to endogenous opiatergic (EOP) neurons. EOP neurons facilitate spontaneous running in hamsters and mediate exercise-induced acceleration of growth and GH pulses. The coincidence of hypoactivity and growth acceleration after such lesions suggested the coexistence of SRIF and EOP fibers within the growth-inhibitory limbic forebrain circuit which control the rate of growth in mature hamsters by the variable inhibition of SRIF neurons by the EOP neurons. This hypothesis posits that accelerated growth is due to increased GH pulse frequency, and hypoactivity due to damage to EOP neurons, and was tested in this study by measuring pulsatile GH release (and as a measure of specificity, pulsatile prolactin release) in the presence and in the absence of opiate-receptor blocker naloxone in 21 female hamsters which sustained electrocoagulative lesions of rostromedial septum and 30 hamsters subjected to control surgery. Lesions doubled GH but not PRL pulse frequency, neither of which was affected by naloxone. Results support the hypothesis that opiatergic neurons facilitate pulsatile GH release by inhibiting the action of somatostatin neurons.

Effects of periventricular lesions on the release of somatostatin during perifusion

Hypothalamic periventricular (PV) nucleus lesions reduce median eminence (ME) SRIF content by ≅ 80% without affecting non-stress plasma growth hormone (GH) levels or the GH response to stress. Our aim was to study the effects of PV lesions on SRIF released during perifusion of preoptic-anterior hypothalamic (PO-AH) tissue. Female rats received anterior or posterior PV lesions; sham-lesioned and intact rats served as controls. Non-stress and stress plasma GH levels were similar in all groups at 2,4 and 16 weeks after surgery. At 18 weeks after surgery, the perifussed PO-AHs of the PV- and sham-lesioned rats released similar amounts of SRIF, and these were higher ( P <0.001) than amounts released from PO-AHs of intact rats. The PO-AHs from all groups showed similar increases in SRIF release after 56 mM K +. Two rats were chosen randomly from each group to asses ME SRIF content; PV lesions caused almost 80% depletion of SRIF, sham lesions did not. These results confirm that most SRIF neurons in the PV nucleus and 80% of ME SRIF content are not essential for the contrl of GH levels under non-stress conditions of for the GH responses to stress and indicate that PV or sham lesions in the rostal forebrain enhance in vitro SRIF release, perhaps from neurons outside the PV nucleus.

Immunocytochemical localization of somatostatin in human brain

The distribution of somatostatin (SRIF) was examined in normal human forebrain, using thick vibratome cut sections. The unlabeled antibody enzyme method of immunocytochemistry revealed a widespread distribution of SRIF immunoreactive neurons and fibers throughout the septum, diencephalon and corpus striatum. Within the septum SRIF neurons and fibers were observed in the medial and lateral septal nuclei, the nucleus of the diagonal band, the nucleus accumbens and the bed nucleus of the stria terminalis. SRIF neurons and fibers were found in several hypothalamic and anterior thalamic nuclei as well as all regions of the corpus striatum. An interesting collection of SRIF immunoreactive neurons and processes were observed forming a wide band extending anteriorly from the lateral preoptic area through the lateral hypothalamus and substantia innominata posteriorly. This report on the localization of immunoreactive SRIF in the human forebrain extends previous anatomical findings and lends morphological support to recent biochemical studies.

The anterior periventricular hypothalamus is the site of somatostatin inhibition of its own release: an in vitro and immunocytochemical study.

The site of action of the inhibitory effect of somatostatin (SRIF) on its own release was studied by: (1) measuring SRIF release in vitro from tissue preparations containing either the proximal (periventricular hypothalamus) or the distal (median eminence) portions of the hypothalamic SRIF neurons, and (2) immunocytochemical investigation of the interconnections occurring between SRIF neuronal elements in these hypothalamic regions. In vitro, a biologically active, but noncross-reacting SRIF analog (D-Trp8 SRIF) in the RIA, inhibited 25 mM K+ induced SRIF release from anterior periventricular hypothalamic tissues. The inhibitory effect of D-Trp8 SRIF was dose-dependent, maximal at 10(-7) M, and restricted to this anterior region, since median-eminence SRIF release was not modified by the presence of D-Trp8 SRIF. Additionally, LHRH release from anterior periventricular hypothalamus was unchanged in the presence of D-Trp8 SRIF. In the periventricular nucleus, perikarya and dendrites of labeled SRIF neurons showed frequent apposition of their limiting membranes. Classical synapses were also observed between SRIF-containing axonal processes and labeled perikarya or dendrites. Although membrane appositions between neighboring SRIF axons frequently occurred in the median eminence, no synaptic-like SRIF-SRIF connections could be detected at this level. The data demonstrate a direct inhibitory action of a SRIF agonist on the anterior periventricular hypothalamic release of the peptide. This effect correlates well with the occurrence of SRIF-SRIF synapses in this region; suggesting that SRIF exerts a negative feedback in the control of its own release through autoreceptors located on the perikarya or dendrites of SRIF-containing neurons.

Long term elevations in plasma thyrotropin, but not growth hormone, concentrations associated with lesion-induced depletion of median eminence somatostatin.

Evidence suggests that somatostatin (SRIF) inhibits nonstress GH and TSH secretion and suppresses GH secretion in response to stress. The aims of this study were to determine whether placement of lesions in the hypothalamic periventricular (PV) nucleus, the location of SRIF neurons which seem to be responsible for most median eminence SRIF, causes elevation of nonstress plasma GH and TSH levels and blocks stress-induced suppression of these hormones. Electrolytic lesions were placed in female rats, and blood was obtained for assessing nonstress and stress plasma levels of GH and TSH at several intervals after surgery until autopsy, 56 weeks after surgery, when median eminences were collected. PV lesions produced only transient elevation of nonstress plasma GH levels and failed to block the suppression of GH secretion by stress. In contrast, PV lesions caused long term elevation of nonstress plasma TSH levels and blockade of stress-induced suppression of TSH secretion. The content and concentration of SRIF in the median eminence were reduced 90% in the PV-lesioned group. These data demonstrate that PV lesions, which result in marked depletion of median eminence SRIF, can cause long term elevations of plasma TSH levels and disruption of the TSH response to stress without producing alterations in GH secretion. Thus, the hypothalamic PV nucleus, its SRIF neurons and most median eminence SRIF are not essential for maintaining normal GH secretion, but seem to be involved in the regulation of TSH release. It appears that different brain structures are involved in inhibiting GH and TSH secretion.

Somatostatin neurons in the rodent hippocampus: An in vitro and in vivo immunocytochemical study

The origin and distribution of immunoreactive-like somatostatin (ISRIF) in the rodent hippocampus was determined by light microscopic immunocytochemistry. ISRIF neurons were demonstrable in long-term explants of fetal mouse hippocampus, demonstrating that a population of SRIF neurons are intrinsic to the hippocampus. In the adult rat, mouse and guinea pig ISRIF was present in several types of neurons - including pyramidal, basket and multipolar neurons - and in all hippocampal regions except for the granule cells of the dentate gyrus. These results suggest that the same molecule is found in neurons which may subserve different functions.

Topography of median eminence somatostatinergic innervation

Somatostatin (SRIF) in the central nervous system is mostly concentrated in the median eminence (ME). Immunocytochemical methods have revealed high densities of SRIF-positive perikarya between the preoptic area and the periventricular nucleus of the hypothalamus (NPE). The aim of the present study was to define more precisely the specific pathways of SRIF neurons from NPE to the ME. SRIF levels were measured by radioimmunoassay, following various hypothalamic transections. Frontal periventricular sections decreased SRIF-ME content by 70% (P less than 0.01), when located at the anterior end of the ME but no diminution was observed when the cuts were located anteriorly or posteriorly. Parasaggital transections decreased SRIF-ME levels by 50% (P less than 0.05) when located at the outer border of the ventromedial and premammillary nucleus, but the decrease was not significant when cuts were located anteriorly. Taken together, our data indicate that most SRIF-containing neurons, originating in the NPE, do not reach the ME directly along the border of the 3rd ventricle; instead they form a loop across the medial forebrain bundle before re-entering the mediobasal hypothalamus at the ME level.

Distribution of somatostatin in the frog brain, Rana catesbiana, in relation to location of catecholamine-containing neuron system.

The distribution of somatostatin (SRIF)-immunoreactive structures in the central nervous system of the bull frog (both with and without treatment of colchicine) was studied, using the indirect immunofluorescence technique of Coons and co-workers (Coons, '58). SRIF-containing cells were observed in more than ten areas including the spinal cord. These SRIF-positive cells showed segmental distribution, in that SRIF-positive neurons were identified in various areas at various brain levels. An extensive network of SRIF-positive fibers was found in most parts of the central nervous system. The distribution of a catecholamine (CA)-containing neuron system in the frog brain is also presented in this study. The possible interactions between SRIF and CA neurons systems are briefly discussed.

Topographic atlas of somatostatin-containing neurons system in the avian brain in relation to catecholamine-containing neurons system. I. Telencephalon and diencephalon.

The morphological organization of the somatostatin (SRIF)-positive neurons in the forebrain (telencephalon and diencephalon) of the warbling grass parakeet (Melopsittacus undulatus) was studied using the indirect immunohistochemical technique of Coons and co-workers ('58). In the telencephalon, a number of SRIF-positive neurons was detected in the lobus paraolfactorius, hippocampus, and paleostriatum. Furthermore, scattered SRIF-labeled cells were noticed in the area corticoidea dorsolateralis and area temporoparieto-occipitalis. A moderate density of immunoreactive fibers was found in the above areas. In addition, although the septal areas was devoid of SRIF-positive neurons, this area contained a moderate occurred in the following hypothalamic areas: (1) nucleus medialis hypothalami posterior, (2) lateral hypothalamus, and (3) mammillary nucleus. The bird hypothalamus also received a strikingly massive SRIF innervation. The heaviest concentration of SRIF-labeled fibers was detected in the medial eminence. Many SRIF-labeled fibers were also observed in other hypothalamic regions. Their locations roughly corresponded in many cases to the areas in which SRIF-positive neurons were disclosed. The overall distribution of the catecholamine system (CA) of the avian forebrain is also represented by means of histofluorescent technique. A possible interaction between SRIF and CA neurons systems is briefly discussed.

Topographic atlas of somatostatin‐containing neuron system in the avian brain in relation to catecholamine‐containing neuron system. II. Mesencephalon, rhombencephalon, and spinal cord

With the indirect immunofluorescence technique of Coon's and collaborators, overall distribution of the somatostatin (SRIF)-positive neurons system in the avian lower brain stem was explored. Numerous cell somata containing SRIF were identified in the interpeduncular nucleus and substantia grisea centralis (GCT) at the level of the nucleus nervi trochlearis. Furthermore, a moderate number of SRIF-positive neurons were seen in the tectum opticum, nucleus tractus solitarii, and spinal cord. Scattered labeled cells were noticed in the rhombencephalon. A dense network of SRIF-positive fibers was distributed widely in the lower brain stem of birds. Their locations corresponded in many cases to the areas where SRIF-positive neurons were found. The present study also presents the distribution of the catecholamine (CA) neuron system in the avian lower brain stem. Possible interactions between SRIF and CA neurons systems are briefly discussed.

Opiate receptors modulate LHRH and SRIF release from mediobasal hypothalamic neurons.

In order to investigate the effect of opiates on luteinizing hormone-releasing hormone (LHRH) and somatostatin (SRIF) release, mediobasal hypothalamic (MBH) slices of male adult rats were superfused at 37 °C with oxygenated Hepes-buffered Locke medium for 1 h. Bacitracin (2 × 10^–5<i>M</i>) was added to prevent enzymatic degradation of neuro-peptides. 6 min pulse of K⁺ (56 m<i>M</i>) markedly stimulated the release of both neuropeptides. Opiates (β-endorphin 10^–7<i>M</i>; D-ALA₂-Met-enkephalin amide 10^–7<i>M</i>; Leuenkephalin 10^–7 M; morphine 10^–6 M) did not alter the spontaneous release of LHRH and SRIF, but significantly inhibited the K⁺-induced neuropeptides release. The effect was reversed by a specific opiate antagonist Naloxone 10^–7<i>M</i>, which was ineffective when perifused alone. β-Endorphin and D-ALA₂-Met-enkephalin inhibited K⁺-evoked LHRH and SRIF release in a dose-dependent manner with a half maximal value of inhibition for concentrations of 7.6 ± 2.7 10^–9 M and 9.4 ± 2.5 10^–9<i>M</i> for LHRH release and 11 ± 2.2 10^–8<i>M</i> and 5.2 ± 3.3 10^–9<i>M</i> for SRIF, respectively. Addition of vasoactive intestinal peptide (VIP) (10-⁹ M) to the medium also inhibited significantly the K⁺-evoked release of SRIF, whereas that of LHRH was not affected. The data suggest: (1) that opiates, acting through specific opiate receptors located on LHRH and SRIF neurons, modulate the release of the neurohormones; (2) the inhibitory effect of opiates could be due to the inhibition of calcium influx through voltage-dependent calcium channels; (3) this interaction may account for the stimulation of growth hormone and the inhibition of luteinizing hormone observed after systemic administration of opiates; (4) VIP inhibits SRIF release, by acting on VIP receptors present on MBH SRIF terminals; the effect is consistent with the stimulation of GH reported after in vivo administration of the peptide.