Renal effects of atriopeptin II and dopamine receptor blockade in acutely volume-expanded rats

Studies were made of the effects of continuous intravenous infusion of a synthetic atrial natriuretic factor (ANF) or, pre-treatment with the dopamine receptor antagonist haloperidol, on the renal response in anaesthetized rats subjected to volume expansion with an isotonic solution at 2% kg-1 body weight (wt) h-1. A time-control group receiving vehicle alone was studied in parallel. Measurements were compared 75 and 145 min after initiation of the volume expansion. Seventy minutes of Atriopeptin II infusion at 10 micrograms h-1 kg-1 body wt did not significantly alter the glomerular filtration rate [control value 1.29 +/- 0.10 ml min-1 g-1 kidney wt (n = 7, mean +/- 1 SEM), experimental value 1.20 +/- 0.12], but increased sodium excretion by 49% (from 2.87 +/- 0.56 to 4.27 +/- 0.45 mumol min-1). The arterial blood pressure was reduced by 9%. In previous investigations we found that in the same dosage Atriopeptin II increased sodium excretion 10-fold in euvolaemic animals. In the time-control group (n = 7) the response was similar to that in the atrial natriuretic factor-treated animals with the exception that the blood pressure was unaltered. Thus, glomerular filtration rate showed no statistically significant change (1.28 +/- 0.06 vs. 1.27 +/- 0.09 ml min-1 g-1 kidney wt) while the sodium excretion increased by 96% (from 2.29 +/- 0.22 to 4.50 +/- 0.49 mumol min-1). In animals pretreated with haloperidol (n = 5), the natriuretic response to the volume expansion was attenuated and was about ten times below that in the time-control group.(ABSTRACT TRUNCATED AT 250 WORDS)     

A possible anti-inflammatory role of angiotensin II type 2 receptor in immune-mediated glomerulonephritis during type 1 receptor blockade

We previously reported that angiotensin II type 1 receptor (AT1R) blockade attenuates renal inflammation/fibrogenesis in immune-mediated glomerulonephritis via angiotensin II type 2 receptor (AT2R). In the present study, further in vivo experiments revealed that AT2R was expressed in tubular epithelial cells of nephritic kidneys in mice, and feedback activation of the renin-angiotensin system during AT1R blockade significantly reduced p-ERK, but not intranuclear nuclear factor-kappaB, levels via AT2R. This led to reduction in mRNA levels of the proinflammatory mediator monocyte chemoattractant protein-1 and overall interstitial inflammation and subsequent fibrogenesis. Specific blockade of ERK expression in tubular epithelium by anti-sense oligodeoxynucleotides also attenuated interstitial inflammation, mimicking the anti-inflammatory action of AT2R in nephritic kidneys. Alternatively, we succeeded in confirming such an AT(2)R function by demonstrating that AT1R blockade did not confer renoprotection in nephritic, AT2R gene-deficient mice. Additional in vitro experiments revealed that AT2R activation did not affect nuclear factor-kappaB activation by tumor necrosis factor-alpha in cultured tubular epithelial cells, although it inhibited ERK phosphorylation, which reduced monocyte chemoattractant protein-1 mRNA levels. These results suggest that feedback activation of AT2Rs in tubular epithelium of nephritic kidneys plays an important role in attenuating interstitial inflammation.     

Effect of central urotensin II on heart rate, blood pressure and brain Fos immunoreactivity in conscious rats

Central administration of urotensin II (UII) increases heart rate (HR), cardiac contractility, and plasma levels of epinephrine and glucose. To investigate the mechanisms causing these responses we examined the effects of i.c.v. administration of rat UII (10 microg) on the sympatho-adrenal and pituitary-adrenal axes in conscious rats, and we mapped the brain sites activated by UII by immunohistochemically detecting Fos expression. In six conscious rats i.c.v. UII, but not vehicle, increased HR significantly 60-90 min after treatment and increased plasma glucose at 60 and 90 min, both indicators of increased epinephrine release. Plasma corticosterone levels were significantly elevated 90 min after i.c.v. UII. Conscious rats, given i.c.v. UII (n=12) and killed after 100 or 160 min, showed increased Fos-immunoreactivity (Fos-IR) in the nucleus of the solitary tract and the central nucleus of the amygdala (CeA) at both time points, compared with vehicle (n=11). In UII-treated rats, Fos-IR in the paraventricular nucleus of the hypothalamus (PVN) was significantly elevated at 160 min, but not 100 min, compared with vehicle. There were no increases in Fos-IR in the rostral ventrolateral medulla or the A5 cell group, areas associated with sympathetic outflow to the adrenal gland. In summary, i.c.v. UII increased HR and plasma glucose and corticosterone in conscious rats. UII increased Fos-IR in the CeA and PVN, but over a longer time course in the latter. These findings indicate that UII acts on specific brain nuclei to stimulate the hypothalamo-pituitary-adrenal axis and to stimulate adrenal sympathetic nerve activity.     

Fos immunoreactivity in some locomotor neural centres of 6OHDA-lesioned rats

In this study, we explore Fos expression (a measure of cell activity) in three nuclei associated with locomotion, namely the zona incerta, pedunculopontine tegmental nucleus and cuneiform nucleus (the latter two form the mesencephalic locomotor region) in hemiparkinsonian rats. Sprague–Dawley rats had small volumes of either saline (control) or 6 hydroxydopamine (6OHDA) injected into the medial forebrain bundle, the major tract carrying dopaminergic nigrostriatal axons. After various post-lesion survival periods, ranging from 2 h to 28 days, rats were perfused with formaldehyde and their brains processed for routine tyrosine hydroxylase and Fos immunocytochemistry. Our results showed a significant increase (P < 0.05) in the number of strongly labelled Fos⁺ cells in the cuneiform nucleus in the 6OHDA-lesioned cases compared to the controls after 7 and 28 days survival periods. By contrast, there were no significant differences (P > 0.05) in the number of strong-labelled Fos⁺ cells in the zona incerta and pedunculopontine nucleus of 6OHDA-lesioned rats compared to controls at any survival period. Many of the Fos⁺ cells within the pedunculopontine and cuneiform nuclei were glutamatergic (35–60%), while none or very few were nitric oxide synthase⁺. In conclusion, we reveal an increase in the number of strongly labelled Fos⁺ cells within the cuneiform nucleus of the so-called defensive locomotive system in 6OHDA-lesioned rats. In relation to Parkinson disease, we suggest that this increase is associated with the akinesia or lack of movement seen in patients.     

Angiotensin II type 1 (AT(1)) receptor blockade enhances the L-NAME-induced vasoconstriction in rat submandibular gland

The vasoregulatory role of nitric oxide (NO) and angiotensin II type 1 (AT(1)) receptors in the circulation of the submandibular gland (SMG) of rats was studied. The glandular blood flow was determined by means of laser Doppler flowmetry and rubidium isotope technique. The data obtained by these two methods correlated well (r = 0.77; P < 0.01). The AT(1) receptor antagonist candesartan (0.5 mg kg(-1), i.v.) reduced the vascular resistance in the SMG by 37 % (P < 0.05). By contrast, the NO synthase blocker L-NAME (15 mg kg(-1), i.v.) significantly increased vascular resistance in the SMG both in candesartan-treated (P < 0.001) and non-treated (P < 0.001) animals. The increase in resistance was greater (P < 0.05) after previous blockade of AT(1) receptors. These findings suggest that the AT(1) receptors have an important role in the vasoregulation of the SMG in the rat. As a result of AT(1) blockade, NO-dependent tone of glandular vessels may be enhanced significantly.     

Angiotensin II AT1 receptor blockade changes expression of renal sodium transporters in rats with chronic renal failure

We aimed to examine the effects of angiotensin II AT(1) receptor blocker on the expression of major renal sodium transporters and aquaporin-2 (AQP2) in rats with chronic renal failure (CRF). During 2 wks after 5/6 nephrectomy or sham operation, both CRF rats (n=10) and sham-operated control rats (n=7) received a fixed amount of low sodium diet and had free access to water. CRF rats (n=10) were divided into two groups which were either candesartan-treated (CRF-C, n=4) or vehicle-treated (CRF-V, n=6). Both CRF-C and CRF-V demonstrated azotemia, decreased GFR, polyuria, and decreased urine osmolality compared with sham-operated rats. When compared with CRF-V, CRF-C was associated with significantly higher BUN levels and lower remnant kidney weight. Semiquantitative immunoblotting demonstrated decreased AQP2 expression in both CRF-C (54% of control levels) and CRF-V (57%), whereas BSC-1 expression was increased in both CRF groups. Particularly, CRF-C was associated with higher BSC-1 expression (611%) compared with CRF-V (289%). In contrast, the expression of NHE3 (25%) and TSC (27%) was decreased in CRF-C, whereas no changes were observed in CRF-V. In conclusion, 1) candesartan treatment in an early phase of CRF is associated with decreased renal hypertrophy and increased BUN level; 2) decreased AQP2 level in CRF is likely to play a role in the decreased urine concentration, and the downregulation is not altered in response to candesartan treatment; 3) candesartan treatment decreases NHE3 and TSC expression; and 4) an increase of BSC-1 is prominent in candesartan-treated CRF rats, which could be associated with the increased delivery of sodium and water to the thick ascending limb.     

Androgen receptor blockade in the MPOA or VMN: effects on male sociosexual behaviors

Previously, our laboratory has shown that androgen receptors in the medial preoptic area (MPOA) and ventromedial nucleus (VMN) are necessary for copulation in male rats. The present study examined whether these receptors are required for other sociosexual behaviors. In Experiment 1, different regions of the VMN were implanted with the antiandrogen hydroxyflutamide (OHF). We found that implants located in anterodorsal portions of the VMN were more effective at inhibiting the restoration of copulation than implants in the posteroventral VMN. In Experiment 2, a second set of male rats was pretested for copulation and other sociosexual behaviors and was castrated. Experimental animals then received Silastic capsules filled with testosterone (T) plus intracranial (IC) implants filled with OHF to selectively block androgen receptors in either the MPOA or VMN. We found that androgen receptor blockade in the MPOA inhibited the restoration of copulation but had no effect on other sociosexual behaviors. OHF directed at the VMN inhibited the restoration of copulation and 50-kHz vocalizations but had no effect on scent marking. Two tests were used to assay sexual motivation: partner preference and conditioned place preference (CPP). Both methods revealed impairments in sexual motivation in the VMN group but not in animals receiving OHF in the MPOA. Taken together, these data suggest that androgen receptors in the MPOA are essential for copulatory performance, while androgen receptors in the VMN are important for copulation, sexual motivation, and androgen-dependent vocalizations.     

Fos immunoreactivity in hypocretin-synthesizing and hypocretin-1 receptor-expressing neurons: effects of diurnal and nocturnal spontaneous waking,stress and hypocretin-1 administration

Hypocretin/orexin modulates sleep-wake state via actions across multiple terminal fields. Within waking, hypocretin may also participate in high-arousal processes, including those associated with stress. The current studies examined the extent to which alterations in neuronal activity, as measured by Fos immunoreactivity, occur within both hypocretin-synthesizing and hypocretin-1 receptor-expressing neurons across varying behavioral state/environmental conditions associated with varying levels of waking and arousal. Double-label immunohistochemistry was used to visualize Fos and either prepro-hypocretin in the lateral hypothalamus or hypocretin-1 receptors in the locus coeruleus and select basal forebrain regions involved in the regulation of behavioral state/arousal. Animals were tested under the following conditions: 1). diurnal sleeping; 2). diurnal spontaneous waking; 3). nocturnal spontaneous waking; and 4). high-arousal waking (diurnal novelty-stress). Additionally, the effects of hypocretin-1 administration (0.07 and 0.7 nmol) on levels of Fos were examined within these two neuronal populations. Time spent awake, scored for the 90-min preceding perfusion, was largely comparable in diurnal spontaneous waking, nocturnal spontaneous waking and high-arousal waking. Nocturnal spontaneous waking and high-arousal waking, but not diurnal spontaneous waking, were associated with increased levels of Fos within hypocretin-synthesizing neurons, relative to diurnal sleeping. Within hypocretin-1 receptor-expressing neurons, only high-arousal waking was associated with increased levels of Fos. Hypocretin-1 administration dose-dependently increased levels of Fos within hypocretin-1 receptor-expressing neurons to levels comparable to, or exceeding, levels observed in high-arousal waking. Combined, these observations support the hypothesis that hypocretin neuronal activity varies across the circadian cycle. Additionally, these data suggest that waking per se may not be associated with increased hypocretin neurotransmission. In contrast, high-arousal states, including stress, appear to be associated with substantially higher rates of hypocretin neurotransmission. Finally, these studies provide further evidence indicating coordinated actions of hypocretin across a variety of arousal-related basal forebrain and brainstem regions in the behavioral state modulatory actions of this peptide system.     

Lidocaine blockade of amygdala output in fear-conditioned rats reduces Fos expression in the ventrolateral periaqueductal gray

We showed recently that conditioned fear to context induces Fos expression in the ventrolateral periaqueductal gray [Neuroscience (1997) 78, 165-177]. Neurons in this region are thought to play an important role in the expression of freezing during conditioned fear. To test the possibility that this activation comes directly from the amygdala, we looked at changes in Fos expression after a unilateral blockade of the ventral amygdalofugal pathway with lidocaine. The pathway contains fibres originating from the central nucleus of the amygdala that project directly and mainly ipsilaterally to the ventrolateral periaqueductal gray. Conditioned fear was evoked by re-exposing rats to the same box in which they had previously received electric footshocks. The test re-exposure was preceded by a unilateral microinjection of lidocaine (2%, 0.5-1 microl; n = 20) or saline (n = 14). Lidocaine was also tested in non-conditioned animals (n = 13). The results show that, when lidocaine was microinjected in the medial part of the central nucleus of the amygdala or along the ventral amygdalofugal pathway of conditioned rats, fear-induced Fos expression in the ventrolateral periaqueductal gray was reduced on the side ipsilateral to the injection (up to 37% reduction in comparison to the contralateral side). Ipsilateral reductions were also observed with saline, but they were weaker (maximum of 27% reduction). Fos expression remained low on both sides in the non-fear-conditioned animals injected with lidocaine. Finally, although freezing was only partly reduced in the conditioned animals unilaterally injected with lidocaine, it was significantly correlated to the ipsilateral reduction in Fos expression. This study provides direct evidence that the projection from the central nucleus of the amygdala to the ventrolateral periaqueductal gray is activated during fear and that it contributes to the Fos response of the ventrolateral periaqueductal gray.     

辛伐他汀对高脂血症大鼠动脉血管紧张素Ⅱ1型受体mRNA表达的下调作用

目的:观察高脂血症时动脉血管紧张素Ⅱ1型受体(AT₁)mRNA表达水平、外周血血管活性物质的变化特点及降脂治疗的作用, 探讨辛伐他汀逆转内皮功能障碍的机制。方法:实验包括正常对照组, 另两组通过4周建立高脂血症模型, 此后继续高脂喂养, 其中辛伐他汀治疗组在高脂喂养同时喂服辛伐他汀10mg·kg^-1·d^-1)而高脂血症组不予药物治疗, 第20周检测3组的血脂变化及观察AT₁mRNA表达水平、外周血AngⅡ和NO浓度。结果:高脂血症组AT₁mRNA表达水平高于、血NO水平低于正常对照组、而AngⅡ和收缩压无显著差异。辛伐他汀治疗组总胆固醇(TC)、甘油三脂(TG)、低密度胆固醇(LDL-C)显著低于高脂血症组, 且动脉组织AT₁mRNA表达水平也显著低于高脂血症组, 血NO含量高于高脂血症组、但血AngⅡ浓度和收缩压未见显著差异。结论:辛伐他汀在调脂的同时, 下调AT₁mRNA的表达、促进一氧化氮的生成, 从而逆转内皮功能障碍、阻止动脉硬化进展。     

Lidocaine blockade of amygdala output in fear-conditioned rats reduces Fos expression in the ventrolateral periaqueductal gray

We showed recently that conditioned fear to context induces Fos expression in the ventrolateral periaqueductal gray [Neuroscience (1997) 78, 165–177]. Neurons in this region are thought to play an important role in the expression of freezing during conditioned fear. To test the possibility that this activation comes directly from the amygdala, we looked at changes in Fos expression after a unilateral blockade of the ventral amygdalofugal pathway with lidocaine. The pathway contains fibres originating from the central nucleus of the amygdala that project directly and mainly ipsilaterally to the ventrolateral periaqueductal gray. Conditioned fear was evoked by re-exposing rats to the same box in which they had previously received electric footshocks. The test re-exposure was preceded by a unilateral microinjection of lidocaine (2%, 0.5–1 μl; n=20) or saline (n=14). Lidocaine was also tested in non-conditioned animals (n=13). The results show that, when lidocaine was microinjected in the medial part of the central nucleus of the amygdala or along the ventral amygdalofugal pathway of conditioned rats, fear-induced Fos expression in the ventrolateral periaqueductal gray was reduced on the side ipsilateral to the injection (up to 37% reduction in comparison to the contralateral side). Ipsilateral reductions were also observed with saline, but they were weaker (maximum of 27% reduction). Fos expression remained low on both sides in the non-fear-conditioned animals injected with lidocaine. Finally, although freezing was only partly reduced in the conditioned animals unilaterally injected with lidocaine, it was significantly correlated to the ipsilateral reduction in Fos expression.This study provides direct evidence that the projection from the central nucleus of the amygdala to the ventrolateral periaqueductal gray is activated during fear and that it contributes to the Fos response of the ventrolateral periaqueductal gray.     

Angiotensin II type 1 receptor blockade partially attenuates hypoxia-induced pulmonary hypertension in newborn piglets: relationship with the nitrergic system

The objective of this study was to observe possible interactions between the renin-angiotensin and nitrergic systems in chronic hypoxia-induced pulmonary hypertension in newborn piglets. Thirteen chronically instrumented newborn piglets (6.3 ± 0.9 days; 2369 ± 491 g) were randomly assigned to receive saline (placebo, P) or the AT₁ receptor (AT₁-R) blocker L-158,809 (L) during 6 days of hypoxia (FiO₂ = 0.12). During hypoxia, pulmonary arterial pressure (Ppa; P < 0.0001), pulmonary vascular resistance (PVR; P < 0.02) and the pulmonary to systemic vascular resistance ratio (PVR/SVR; P < 0.05) were significantly attenuated in the L (N = 7) group compared to the P group (N = 6). Western blot analysis of lung proteins showed a significant decrease of endothelial NOS (eNOS) in both P and L animals, and of AT₁-R in P animals during hypoxia compared to normoxic animals (C group, N = 5; P < 0.01 for all groups). AT₁-R tended to decrease in L animals. Inducible NOS (iNOS) did not differ among P, L, and C animals and iNOS immunohistochemical staining in macrophages was significantly more intense in L than in P animals (P < 0.01). The vascular endothelium showed moderate or strong eNOS and AT₁-R staining. Macrophages and pneumocytes showed moderate or strong iNOS and AT₁-R staining, but C animals showed weak iNOS and AT₁-R staining. Macrophages of L and P animals showed moderate and weak AT₂-R staining, respectively, but the endothelium of all groups only showed weak staining. In conclusion, pulmonary hypertension induced by chronic hypoxia in newborn piglets is partially attenuated by AT₁-R blockade. We suggest that AT₁-R blockade might act through AT₂-R and/or Mas receptors and the nitrergic system in the lungs of hypoxemic newborn piglets.     

Neonatal capsaicin treatment decreased substance P receptor immunoreactivity in lamina III neurons of the dorsal horn

Using immunohistochemistry and in situ hybridization technique, the distribution of substance P (SP) and SP receptors was studied in the dorsal horn of the rat spinal cord after neonatal capsaicin treatment. Sprague-Dawley rats administered 100 mg/kg of capsaicin subcutaneously within 24 h after birth were examined at 8 weeks of age. In the capsaicin administered rats, slight reduction of SP immunoreactivities in lamina I, and severe decrease in lamina II were observed. In the control group, SP receptor-mRNA was observed in all laminae, and SP receptor-immunoreactivities were seen to be intense in laminae I and III. In contrast, in the capsaicin administered rats, the SP receptor-mRNA expression was low in laminae II-V, and SP receptor immunoreactivities decreased in laminae III-V. Furthermore, the density of the SP receptor immunoreactivities was considerably decreased in the nerve cells of lamina III. We concluded that elevation of the threshold to painful stimulation in rats was as a result of the decrease in SP immunoreactive afferent fibers in laminae I and II, decrease of the SP receptor-mRNA in the laminae II-V, or the decrease in SP receptor immunoreactive neurons in laminae III-V.     

Posttraining D1 receptor blockade impairs odor conditioning in neonatal rats.

Rat pups that were exposed to a novel anise odor paired with tactile stimulation (stroking the skin with a paint brush) received injections of either saline or the dopamine D1 receptor antagonist (+/-)-SKF 83566 (0.1 mg/kg) before conditioning or immediately after conditioning. Animals that received the drug either before or after training showed less approach to the conditioned odor during the testing period 24 hr later than did animals that received the vehicle. Posttraining administration of the D2 receptor antagonist spiperone (0.1 mg/kg) did not affect subsequent approach to the conditioned odor, suggesting a selective effect of D1 receptor blockade. The impairment in learning by the administration of (+/-)-SKF 83566 before conditioning was reversed by the injection of the dopamine receptor agonist apomorphine (0.1 mg/kg) immediately after conditioning. Posttraining D1 receptor activation appears necessary for normal odor conditioning in rat pups.     

Inhibition of drinking in water-deprived rats by combined central angiotensin II and cholinergic receptor blockade.

The effect of blockade of central angiotensin II (AII) receptors and cholinergic receptors on thirst induced by water deprivation was studied in Sprague-Dawley rats and rats with hereditary hypothalamic diabetes insipidus (DI). Neither central AII nor cholinergic blockade alone affected drinking. Antagonism of both receptors simultaneously, however, significantly inhibited water intake of both Sprague-Dawley and DI rats. This inhibitory effect was not observed in water-deprived, nephrectomized rats. The combined antagonism on water intake was specific, since milk intake in hungry rats was not affected by simultaneous AII and cholinergic blockade. Isorenin concentrations in brain tissue were at control levels in water-deprived, nephrectomized, and non-nephrectomized Sprague-Dawley rats but were increased in water-deprived DI rats. The results suggest that angiotensin and cholinergic receptors in the brain have a physiological role in thirst. Thirst is maintained when either receptor is intact, but reduced when both receptors are inhibited by antagonists. They are independently capable of maintaining thirst.     

Distribution of androgen receptor immunoreactivity in the brainstem of male rats

Gonadal steroids such as testosterone and estrogen are necessary for the normal activation of male rat sexual behavior. The medial preoptic area (MPOA), an important neural substrate regulating mating, accumulates steroids and also expresses functional androgen receptors (AR). The MPOA is intimately connected with other regions implicated in copulation, such as the bed nucleus of the stria terminalis and medial amygdala. Inputs to the MPOA arise from several areas within the brainstem, synapsing preferentially onto steroid sensitive MPOA cells which are activated during sexual activity. Given that little is known about the distribution of AR protein in the brainstem of male rats, we mapped the distribution of AR expressing cells in the pons and medulla using immunocytochemistry. In agreement with previous reports, AR immunoreactivity (AR-ir) was detected in ventral spinal motoneurons and interneurons. In addition, AR-ir was detected in areas corresponding to the solitary tract, lateral paragigantocellular and alpha and ventral divisions of the gigantocellular reticular nuclei, area postrema, raphe pallidus, ambiguus nucleus, and intermediate reticular nucleus. Several regions within the pons contained AR-ir, such as the tegmental and central gray, parabrachial nucleus, locus coeruleus, Barrington's nucleus, periaqueductal gray, and dorsal raphe. In contrast with in situ hybridization studies, auditory and somatosensory areas were AR-ir negative, and, except for very light staining in the prepositus nucleus, areas carrying vestibular information did not display AR-ir. Additionally, cranial nerve motoneurons of the hypoglossal, facial, dorsal vagus, and spinal trigeminal did not display AR-ir in contrast to previous reports. The data presented here indicate that androgens may influence numerous cell groups within the brainstem. Some of these probably constitute a steroid sensitive circuit linking the MPOA to motoneurons in the spinal cord via androgen responsive cells in the caudal ventral medulla.