Participation of galaninergic neurotransmission at the paraventricular hypothalamic nucleus in the suppression of baroreceptor reflex response by locus ceruleus in the rat

We evaluated the potential participation of galanin (GAL) at the paraventricular nucleus of hypothalamus (PVN) in the suppression of baroreceptor reflex (BRR) response by locus ceruleus (LC), using adult male Sprague-Dawley rats anesthetized with pentobarbital sodium. Microinjection of GAL (100 pmol) bilaterally into the PVN significantly depressed the BRR response. This suppressive effect was appreciably antagonized when GAL (100 pmol) and GAL antiserum (1:20) were coadministered into the bilateral PVN. Whereas bilateral microinjection of GAL antiserum into the PVN by itself elicited minimal effect, it nevertheless significantly attenuated the suppressive effect of either electrical or chemical activation of LC on the BRR response. Pretreatment with the same amount of normal rabbit serum (1:20), on the other hand, was ineffective. These results suggest that a galaninergic projection from the LC to PVN may participate in the suppression of BRR response by this dorsal pontine nucleus.     

Studies on the presence of angiotensin II in rat brain

Abstract: Angiotensin II-like immunoreactivity was extracted from brains of bilaterally nephrectomized rats with several different extraction procedures (90% methanol, distilled water, 6 M urea, 0.1 N HCI, and 2 M acetic acid). The activity was measured with radioimmunoassays using three different antisera, two of which had been used previously for immunocytochemical studies. With none of the extraction procedures or antisera employed was more than 80 pg/g wet weight of angiotensin II-like immunoreactivity found. Analysis was undertaken with two different reverse-phase high-pressure liquid chromatography systems; in one of these the immunoreactivity did not coelute with angiotensin II or III. On the basis of its elution pattern from a molecular sieving column, the immunoreactivity seems to have a higher molecular weight than angiotensin II. It is concluded that neurons in the brain do not synthesize and store angiotensin II.     

室旁核注射GLP-1 及拮抗剂对大鼠胃组织nesfatin-1 表达的影响

目的:探讨下丘脑室旁核(pareventricular,PVN)注射胰高血糖素样肽-1(GLP-1)及其受体拮抗剂Exendin(9-39)后胃组织核组蛋白2(NUCB2)/nesfatin-1表达的影响。方法:选取48只雄性Wistar大鼠,随机分为6组,生理盐水组,四种不同剂量GLP-1组(0.003 nmol/10μL,0.03 nmol/10μL,0.3 nmol/10μL,3 nmol/10μL),30 nmol Exendin(9-39)+3 nmol GLP-1(E+G)组,每组8只。PVN区埋置套管并按每组要求分别经套管给予GLP-1及Exendin(9-39)等药物。给药2小时后处死大鼠并取胃组织,实时荧光定量RT-PCR法检测各组胃组织NUCB2 m RNA表达。另外生理盐水组,3 nmo L GLP-1组及E+G组每组分别随机取6只大鼠的部分胃组织,用免疫组织化学法测胃粘膜NUCB2/nesfatin-1蛋白的表达情况。结果:实时荧光定量RT-PCR法发现3 nmo L GLP-1组大鼠胃组织NUCB2 m RNA表达量高于生理盐水组,差异有统计学意义(P0.05),而其余各组大鼠胃组织NUCB2 m RNA表达与生理盐水组比较无统计学差异(P0.05)。免疫组化结果显示3 nmo L GLP-1组胃粘膜NUCB2/nesfatin-1蛋白表达与生理盐水组、E+G组比较有统计学差异(P0.05),生理盐水组大鼠胃粘膜NUCB2/nesfatin-1蛋白表达与E+G组比较无明显差异(P0.05)。结论:PVN注射GLP-1能够促进胃组织NUCB2/nesfatin-1的表达,这一作用可能是通过激活GLP-1受体来完成的。     

Immunofluorescence studies on gonadotropin releasing hormone (GRH) in the fore-brain and the neurohypophysis of the green frog,Rana esculenta L.

Summary Using antibodies against mammalian LH-RH, the double antibody immunofluorescence technique has been applied to serial cross sections of the brains of adult Rana esculenta. Immunoreactive material was found in perikarya of an unpaired nucleus in front of the preoptic recess. The axons of these perikarya also contain fluorescing material. They form a single bundle which passes under the preoptic recess, than splits into two tracts, one on either side of the optic chiasm. The two tracts reunite just before entering the median eminence. The axons end near the capillaries in the outer zone of the median eminence. The possibility of two separate centres for the stimulation of gonadotropic activity in the brains of anurans is discussed.The authors wish to thank Mr. E. von der Vlist and Mr. J.J. van der Vlis for preparing the illustrations     

In vivo imaging of oxidative stress in the kidney of diabetic mice and its normalization by angiotensin II type 1 receptor blocker

This study was undertaken to evaluate oxidative stress in the kidney of diabetic mice by electron spin resonance (ESR) imaging technique. Oxidative stress in the kidney was evaluated as organ-specific reducing activity with the signal decay rates of carbamoyl-PROXYL probe using ESR imaging. The signal decay rates were significantly faster in corresponding image pixels of the kidneys of streptozotocin-induced diabetic mice than in those of controls. This technique further demonstrated that administration of angiotensin II type 1 receptor blocker (ARB), olmesartan (5 mg/kg), completely restored the signal decay rates in the diabetic kidneys to control values. In conclusion, this study provided for the first time the in vivo evidence for increased oxidative stress in the kidneys of diabetic mice and its normalization by ARB as evaluated by ESR imaging. This technique would be useful as a means of further elucidating the role of oxidative stress in diabetic nephropathy.     

Intrarenal localization of angiotensin II specific binding in rat fetuses

Specific binding sites for angiotensin II were localized in the developing rat kidney (18th day of pregnancy and immediately before birth) by autoradiography using [125I]-ileu-5-angiotensin II either perfused in vivo through the fetal aorta or added in vitro to frozen sections in an incubation mixture. Specific binding was localized in the walls of the afferent and efferent arterioles, in the intraglomerular cells and in the peritubular arterioles of the subcapsular cortical zone. The immunohistochemical analysis, carried out on receptors saturated with unlabelled angiotensin II perfused through the mother's aorta, confirmed the autoradiographical localization. Antisera against ileu-5-angiotensin II were used in the indirect immunofluorescence technique and in the PAP method. Immunolocalization of angiotensin II was also found in the proximal tubule and in the thick ascending limb of Henle's loop.     

Golgi-type and immunocytochemical studies on the intrinsic organization of the periventricular layer of the avian paraventricular nucleus

Summary By means of the rapid Golgi-impregnation technique, intraependymal nerve cells and CSF-contacting neurons were demonstrated in the periventricular layer of the paraventricular nucleus (PVN) of the Japanese quail, chicken and house sparrow. When immunocytochemistry was applied to the brains of Japanese quail, chicken and Pekin duck these cellular elements displayed vasotocin- and neurophysin immunoreactions. In the present material intraependymal and cerebrospinal fluid-contacting neurons of the paraventricular nucleus (PVN) were never stained with antisera against avian vasointestinal peptide (VIP), corticotropin releasing factor (CRF), angiotensin II and serotonin. The periventricular zone of the PVN comprises conspicuous fiber networks immunoreactive with antisera to neurophysin, vasotocin and CRF. Our results indicate a general morphologic pattern of the above-mentioned peptidergic systems in all avian species investigated, irrespective of their taxonomic position or of certain peculiarities of the habitat and functional adaptation. The present neuromorphological results may provide a basis for further functional analysis of the interrelationships between the compartments containing cerebrospinal fluid and the neuroendocrine hypothalamic centers involved in various autonomic control mechanisms.Supported by grants from the MPI (60%), the Italian CNR (83.00447.04, 84.01769.04, 85.00515.04), and the Deutscher Akademischer Austauschdienst to G.C.P. and C.V.P., and from the Deutsche Forschungsgemeinschaft to H.W.K. (Ko 758/2-4)Preliminary results were previously presented at the 78th Versammlung der Anatomischen Gesellschaft (Viglietti-Panzica et al. 1984a) and at the 8th meeting of the European Neuroscience Association (Viglietti-Panzica et al. 1984b)     

Heterogeneity of angiotensin II receptors in membranes of developing rat metanephros

Specific and high affinity binding sites for angiotensin II were demonstrated in the membranes of the developing rat metanephros during the second half of pregnancy and in the newborn by binding studies with 125I angiotensin II. Only one type of angiotensin receptor was found during intrauterine life while after birth two classes of angiotensin receptors were present in the membranes of the cortical renal tissue.     

Involvement of endogenous ouabain-like compound in the cardiac hypertrophic process in vivo

The Na(+)/K(+)-ATPase inhibitor ouabain has been shown to trigger hypertrophic growth of cultured cardiomyocytes; however, the significance of endogenous ouabain-like compound (OLC) in the hypertrophic process in vivo is unknown. Here we characterized the involvement of OLC in left ventricular (LV) hypertrophy induced by norepinephrine (NE) and angiotensin II (Ang II) infusions in rats. Administration of NE (300 microg/kg/h) via subcutanously implanted osmotic minipumps for 72 h resulted in a significant increase in left ventricular weight to body weight (LVW/BW) ratio (P<0.001) and a substantial up-regulation of atrial natriuretic peptide (ANP) gene expression (13.2-fold, P<0.001). NE infusion induced a transient increase in plasma OLC levels at 12 h (P<0.05), which returned to control levels by 72 h. Adrenalectomy markedly reduced both basal and NE-induced increase in plasma OLC levels. LVW/BW ratio was not modulated by adrenalectomy; however, ANP gene expression was blunted by 44% (P<0.01) and 47% (P<0.05) at 12 and 72 h, respectively. In agreement, adrenalectomy reduced up-regulation of ANP without affecting LV mass in rats infused with Ang II (33 microg/kg/h). Administration of exogenous ouabain (1 nM to 100 microM) for 24 h had no effect on ANP gene expression in cultured neonatal rat ventricular myocytes. However, the up-regulation of ANP mRNA levels induced by the alpha-adrenergic agonist phenylephrine (1 microM) was markedly enhanced by ouabain (100 microM) (5.6-fold vs. 9.6-fold, P<0.01). These data show that OLC as an adrenal-derived factor may be required for the induction LV ANP gene expression during the hypertrophic process.     

The topography of the caudal part of the paraventricular organ in Rana temporaria

Summary The results of a monoamine-fluorescence study of the hypothalamus of Rana temporaria show that the brain area corresponding with the nucleus infundibularis dorsalis (NID), as described in other species, does not differ, neither morphologically nor histochemically, from the paraventricular organ (PVO), with which it is anatomically continuous. It is concluded that a nucleus infundibularis dorsalis does not exist as a separate entity in this species.     

Angiotensins II and IV stimulate the activity of tyrosine kinases in estrogen-induced rat pituitary tumors

The effects of angiotensin II (AngII) and its fragment 3-8 (angiotensin IV, AngIV) on tyrosine kinase (TK) activity in estrogen-induced rat pituitary tumor homogenates were studied. It was found that both angiotensin peptides increase the TK activity. AngIV was effective in a lower concentration and its maximal effect was greater as compared to that of AngII. Moreover, the specific inhibitors of aminopeptidase A (EC33) and of aminopeptidase N (PC18) significantly attenuated the stimulatory effect of AngII. Because the action of both aminopeptidases is necessary to convert AngII into AngIV, this finding suggested that the effect of AngII on TK activity is (at least in part) exerted via AngIV.     

Immunocytochemical localization of angiotensin-(1–7) in the rat forebrain

Recent findings by this group have led us to reconsider the view that amino (N−) terminal fragments of angiotensin (Ang) II are inactive degradation products of renin-angiotensin system. To further examine this possibility, an antibody to Ang-(1–7), the N-terminal heptapeptide, was produced to demonstrate the neuroanatomical distribution of the rat brain. Ang-(1–7)-immunoreactivity was found in paraventricular, supraoptic, and suprachiasmatic nuclei, bed nucleus of the stria terminalis, substantia innominata, median eminence, and neurohypophysis. This distribution of Ang-(1–7) in the rat forebrain, together with our previous demonstrations of vasopressin secretion in response to this peptide, suggest that Ang-(1–7) functions as a neuromodulator.     

Parabrachial nucleus induces suppression of baroreflex bradycardia by the release of glutamate in the rostral ventrolateral medulla of the rat

The involvement of glutamatergic neurotransmission in the rostral ventrolateral medulla (RVLM) in the suppression of baroreflex bradycardia by the parabrachial nucleus (PBN) was investigated. Repeated electrical activation of the PBN increased the concentration of glutamate in the dialysate collected from the RVLM. The same stimulation also suppressed baroreflex bradycardia in response to transient hypertension evoked by phenylephrine (5 microg/kg, intravenously). Microinfusion of L-glutamate (10, 50 or 100 microM) via the microdialysis probe into the RVLM dose-dependently elicited a significant inhibition of baroreflex bradycardia that paralleled the concentration and time course of the PBN-elicited elevation in extracellular glutamate in the RVLM. The suppression of baroreflex bradycardia elicited by microinjection of L-glutamate (1 nmol) into the RVLM was appreciably reversed by coinjection of the NMDA receptor antagonist, dizocilpine (500 pmol), or the non-NMDA receptor antagonist, 6-cyano-7-nitroquinoxaline-2, 3-dione (50 pmol). These results suggest that an increase in the extracellular concentration of glutamate and activation of both NMDA and non-NMDA receptors in the RVLM may mediate the suppression of baroreflex bradycardia by activation of the PBN.     

Effect of chronic ethanol consumption on emotional stress in the white rat

Chronic pain emotional stress (PES), paired action of the white noise and electric skin stimulation and chronic (during 7 months) ethanol consumption in white rats were shown to act in the same direction. Hypertension, decrease of respiratory rate and increase of Hildebrandt index were observed as a result of PES, ethanol consumption, and especially under PES during ethanol consumption. Ethanol consumption by the animals led to their growth retardation and increase of the spleen and heart mass. Accidental thymus involution was noted both under ethanol consumption and PES. Activation of lipid peroxidation and decrease of superoxide dismutase activity (of its mitochondrial form especially) as well as of Na+,K+-ATP-ase activity were observed in brain homogenates of the rats after PES, while the general ATP-ase activity remained unchanged. An increase of triiodothyronine level and the tendency to thyroxine level increase as well as a decrease of superoxide dismutase activity were observed in the blood serum of these animals. A tendency towards lipid peroxidation level decrease and to brain superoxide dismutase activity increase, as well as blood antioxidation activity increase (evaluated by transferrin and coeruloplasmin contents and by serum superoxide dismutase activity) and a decrease of thyroxine level were observed as a result of ethanol consumption. The mechanisms are discussed of the "anti-stress" action of short-term ethanol consumption and of the action of its chronic consumption, additive to PES.     

Effect of clonidine on cardiac baroreflex delay in humans and rats

The delay τ between rising systolic blood pressure (SBP) and baroreflex bradycardia has been found to increase when vagal tone is low. The α(2)-agonist clonidine increases cardiac vagal tone, and this study tested how it affects τ. In eight conscious supine human volunteers clonidine (6 μg/kg po) reduced τ, assessed both by cross correlation baroreflex sensitivity and sequence methods (both P < 0.05). Experiments on urethane-anaesthetized rats reproduced the phenomenon and investigated the underlying mechanism. Heart rate (HR) responses to increasing SBP produced with an arterial balloon catheter showed reduced τ (P < 0.05) after clonidine (100 μg/kg iv). The central latency of the reflex was unaltered, however, as shown by the unchanged timing with which antidromically identified cardiac vagal motoneurons (CVM) responded to the arterial pulse. Testing the latency of the HR response to brief electrical stimuli to the right vagus showed that this was also unchanged by clonidine. Nevertheless, vagal stimuli delivered at a fixed time in the cardiac cycle (triggered from the ECG R-wave) slowed HR with a 1-beat delay in the baseline state but a 0-beat delay after clonidine (n = 5, P < 0.05). This was because clonidine lengthened the diastolic period, allowing the vagal volleys to arrive at the heart just in time to postpone the next beat. Calculations indicate that naturally generated CVM volleys in both humans and rats arrive around this critical time. Clonidine thus reduces τ not by changing central or efferent latencies but simply by slowing the heart.     

Influence of sarmesin on the cardiac and vascular actions of angiotensin II

Summary The angiotensin II (ANG II) receptor blocker properties of sarmesin and its influence on the homotropic cooperativity of ANG II receptors were studied in two experimental models: isolated rabbit aorta and isolated rabbit atria. The results show that: (i) sarmesin is a specific competitive antagonist of ANG II receptors, with high affinity (pA₂=8.93 in the isolated aorta and 8.66 in the isolated atria); and (ii) the slope of the concentration-response curves to ANG II and the Hill coefficient increased in the presence of sarmesin, the latter suggesting an enhancement of the positive homotropic cooperativity of ANG II receptors. These results may be explained overall by the reciprocal negative modulation of receptor affinity between sarmesin and ANG II, due, possibly, to opposite effects on the binding of G-proteins to ANG II receptors.     

Involvement of calcium-sensing receptor in cardiac hypertrophy-induced by angiotensinII through calcineurin pathway in cultured neonatal rat cardiomyocytes

Cardiac hypertrophy is a common pathological change accompanying cardiovascular disease. Recently, some evidence indicated that calcium-sensing receptor (CaSR) expressed in the cardiovascular tissue. However, the functional involvement of CaSR in cardiac hypertrophy remains unclear. Previous studies have shown that CaSR caused accumulation of inositol phosphate to increase the release of intracellular calcium. Moreover, Ca²⁺-dependent phosphatase calcineurin (CaN) played a vital role in the development of cardiac hypertrophy. Therefore, we investigated the expression of CaSR in cardiac hypertrophy-induced by angiotensin II (AngII) and the effects of CaSR activated by GdCl₃ on the related signaling transduction pathways. The results showed that AngII induced cardiac hypertrophy and up-regulated the expression of CaSR, meanwhile increased the intracellular calcium concentration ([Ca²⁺]_i) and activated CaN hypertrophic signaling pathway. Compared with AngII alone, the above changes were further obvious when adding GdCl₃. But the effects of GdCl₃ on the cardiac hypertrophy were attenuated by CsA, a specific inhibitor of CaN. In conclusion, these results suggest that CaSR is involved in cardiac hypertrophy-induced by AngII through CaN pathway in cultured neonatal rat cardiomyocytes.     

Angiotensin II and norepinephrine antagonize the secretory effect of VIP in rat ileum and colon

Vasoactive intestinal polypeptide (VIP) induces intestinal secretion of water and electrolytes in experimental animals and man. We assessed the ability of angiotensin II (AII) and norepinephrine (NE) to block the secretion evoked by VIP, in vivo. Ileal and colonic segments in rats were perfused in situ for two hours with a physiological buffer containing [14C]-PEG-4000 as a volume marker. Saline (0.9% NaCl) was infused intravenously during the first hour and VIP or a combination of VIP plus AII or NE was infused during the second hour. All (0.7 ng/kg/min) alone enhanced water absorption significantly (p less than 0.01) in the ileum and an appreciable, although not a statistically significant, effect was observed in the colon. AII antagonized the secretory effects of VIP in the ileum as well as in the colon. Norepinephrine (5 micrograms/kg/min) also reversed the effect of VIP on the small intestine and colon. Although the mechanism by which AII antagonizes the secretory effects of VIP has not been identified, it is probable that AII promotes absorption, at least in part secondary to release of mucosal NE.     

In vitro autoradiographic localization of [I]-angiotensin II binding sites in the rat and dog kidney

Light microscopic autoradiographic techniques have been utilized to demonstrate specific regions of the rat and dog kidney where angiotensin II receptors exist. Slide mounted tissue sections were labeled with [¹²⁵I]-angiotensin II using conditions which provided for highly specific binding. These angiotensin II binding sites were localized to several distinct renal structures. In the renal cortex, angiotensin II binding sites were found concentrated in all parts of the glomeruli including the vascular components, the macula densa and the juxtaglomerular apparatus. Angiotensin II binding in the medulla was more diffusely associated with the vasa recta, and to a lesser extent, the thick ascending segment of the loop of Henle. Binding sites specific for angiotensin II were also found in the smooth muscle laminae of the ureter. Scatchard analysis of the binding kinetics allowed the demonstration of two subpopulations of binding sites which differ slightly in their affinities for [¹²⁵I]-angiotensin II. These subpopulations appear to be associated with distinct components of the renal structure.     

Losartan, an antagonist of AT1 receptor for angiotensin II, attenuates lipopolysaccharide-induced acute lung injury in rat

Angiotensin II (Ang II) plays an important role in inflammatory process. Acute lung injury (ALI), an inflammatory disorder of the lung, is commonly associated with endotoxemia; however, the mechanism that endotoxin (lipopolysaccharide, LPS) induces the inflammatory response in ALI is not well defined. Here, we showed, in LPS-induced ALI rat model, that Ang II and Ang II type 1 (AT₁) receptor were significantly increased in lung tissues, compared with those in controls. Meanwhile, nuclear factor (NF)-κB-DNA-binding activity, tumor necrosis factor (TNF)-α mRNA, and pneumocytic apoptosis were significantly increased. Moreover, pretreatment of rats with losartan, an antagonist of AT₁ receptor for Ang II, improved the inflammation, reduced the elevation of Ang II and AT₁ receptor, and inhibited NF-κB-DNA-binding activity, expression of TNF-α mRNA, and pneumocytic apoptosis. The data indicate that Ang II may mediate the inflammatory process in LPS-induced ALI through AT₁ receptor, which can be blocked by losartan.