Mineralocorticoid Receptor Antagonist RU28318 Research Articles

Acute hypernatremia dampens stress-induced enhancement of long-term potentiation in the dentate gyrus of rat hippocampus.

Stress often occurs within the context of homeostatic threat, requiring integration of physiological and psychological demands to trigger appropriate behavioral, autonomic and endocrine responses. However, the neural mechanism underlying stress integration remains elusive. Using an acute hypernatremic challenge (2.0M NaCl subcutaneous), we assessed whether physical state may affect subsequent responsiveness to psychogenic stressors. We found that experienced forced swimming (FS, 15min in 25°C), a model of psychogenic stress, enhanced long-term potentiation (LTP) induction in the dentate gyrus (DG) of the rat hippocampus ex vivo. The effect of FS on LTP was prevented when the animals were adrenalectomized or given mineralocorticoid receptor antagonist RU28318 before experiencing stress. Intriguingly, relative to normonatremic controls, hypernatremic challenge effectively elevated plasma sodium concentration and dampened FS-induced enhancement of LTP, which was prevented by adrenalectomy. In addition, acute hypernatremic challenge resulted in increased extracellular signal-regulated kinase (ERK)1/2 phosphorylation in the DG and occluded the subsequent activation of ERK1/2 by FS. Moreover, stress response dampening effects by acute hypernatremic challenge remained intact in conditional oxytocin receptor knockout mice. These results suggest that acute hypernatremic challenge evokes a sustained increase in plasma corticosterone concentration, which in turn produces stress-like changes in the DG, thereby occluding subsequent responsiveness to psychogenetic stress. They also fit into the general concept of "metaplasticity" - that is, the responsiveness to stress is not fixed but appears to be governed by the recent history of prior physical state.

Deletion of the forebrain mineralocorticoid receptor impairs social discrimination and decision-making in male, but not in female mice

Social interaction with unknown individuals requires fast processing of information to decide whether it is friend or foe. This process of discrimination and decision-making is stressful and triggers secretion of corticosterone activating mineralocorticoid receptor (MR) and glucocorticoid receptor (GR). The MR is involved in appraisal of novel experiences and risk assessment. Recently, we have demonstrated in a dual-solution memory task that MR plays a role in the early stage of information processing and decision-making. Here we examined social approach and social discrimination in male and female mice lacking MR from hippocampal-amygdala-prefrontal circuitry and controls. The social approach task allows the assessment of time spent with an unfamiliar mouse and the ability to discriminate between familiar and unfamiliar conspecifics. The male and female test mice were both more interested in the social than the non-social experience and deletion of their limbic MR increased the time spent with an unfamiliar mouse. Unlike controls, the male MRCaMKCre mice were not able to discriminate between an unfamiliar and the familiar mouse. However, the female MR mutant had retained the discriminative ability between unfamiliar and familiar mice. Administration of the MR antagonist RU28318 to male mice supported the role of the MR in the discrimination between an unfamiliar mouse and a non-social stimulus. No effect was found with a GR antagonist. Our findings suggest that MR is involved in sociability and social discrimination in a sex-specific manner through inhibitory control exerted putatively via limbic-hippocampal efferents. The ability to discriminate between familiar and unfamiliar conspecifics is of uttermost importance for territorial defense and depends on a role of MR in decision-making.

Abstract 415: Brain Epidermal Growth Factor Receptor and c-Src Tyrosine Kinase Contribute to Sympathetic Excitation Induced by Systemically Administered Aldosterone in Rats

Introduction: Epidermal growth factor receptor (EGFR) and c-Src tyrosine kinase mediate multiple functions of aldosterone and angiotensin II in peripheral cardiovascular tissues by regulating mitogen-activated protein kinase (MAPK) signaling. Hypothesis: EGFR and c-Src activity in cardiovascular regions of the brain contribute to sympatho-excitatory responses induced by circulating aldosterone. METHODS: In anesthetized rats, we recorded renal sympathetic nerve activity (RSNA), mean blood pressure (MBP) and heart rate (HR) at baseline and for 4 hours during intravenous infusion of aldosterone (30 μg/hr), with or without concomitant intracerebroventricular (ICV) infusion of the EGFR inhibitor AG1478 (1 nmol/hr), the mineralocorticoid receptor (MR) antagonist RU28318 (10 μg/hr) or the p44/42 MAPK inhibitor PD98059 (5 μg/hr), or pretreatment with a bolus ICV injection of the c-Src inhibitor SU6656 (1 nmol). In some rats, hypothalamic paraventricular nucleus (PVN) tissues were collected to examine the expression of phosphorylated p44/42 MAPK. Results: Aldosterone infusion significantly (*p<0.05 vs baseline) increased RSNA (25.3 ± 4.6 %*), MBP (9.8 ± 2.2 mmHg*) and HR (28.2 ± 6.2 bpm*), as well as the PVN expression of phosphorylated p44/42 MAPK (86%, p<0.05), as compared with vehicle. Concomitant ICV infusion of the EGFR inhibitor AG1478 significantly (†p<0.05 vs aldosterone alone) diminished the phosphorylation of p44/42 MAPK (28 %†) in PVN and the increases in RSNA (3.6 ± 1.5 %†), MBP (3.2 ± 1.3 mmHg†) and HR (5.3 ± 5.1 bpm†) induced by aldosterone. ICV pretreatment with the c-Src inhibitor SU6656 had similar significant (p<0.05) inhibitory effects on the RSNA, MBP and HR responses to aldosterone. Concomitant ICV infusion of either the MR antagonist RU28318 or the p44/42 MAPK inhibitor PD98059 prevented the aldosterone-induced sympathetic excitation. Conclusion: Central inhibition of EGFR and c-Src significantly reduced the brain MR- and MAPK-dependent sympathetic excitation elicited by systemically administered aldosterone. Brain EGFR and c-Src are potential therapeutic targets for cardiovascular disease states characterized by high circulating levels of aldosterone.

Aldosterone-induced brain MAPK signaling and sympathetic excitation are angiotensin II type-1 receptor dependent

Angiotensin II (ANG II)-induced mitogen-activated protein kinase (MAPK) signaling upregulates angiotensin II type-1 receptors (AT(1)R) in hypothalamic paraventricular nucleus (PVN) and contributes to AT(1)R-mediated sympathetic excitation in heart failure. Aldosterone has similar effects to increase AT(1)R expression in the PVN and sympathetic drive. The present study was undertaken to determine whether aldosterone also activates the sympathetic nervous system via MAPK signaling and, if so, whether its effect is independent of ANG II and AT(1)R. In anesthetized rats, a 4-h intravenous infusion of aldosterone induced increases (P < 0.05) in phosphorylated (p-) p44/42 MAPK in PVN, PVN neuronal excitation, renal sympathetic nerve activity (RSNA), mean blood pressure (MBP), and heart rate (HR). Intracerebroventricular or bilateral PVN microinjection of the p44/42 MAPK inhibitor PD-98059 reduced the aldosterone-induced RSNA, HR, and MBP responses. Intracerebroventricular pretreatment (5 days earlier) with pooled small interfering RNAs targeting p44/42 MAPK reduced total and p-p44/42 MAPK, aldosterone-induced c-Fos expression in the PVN, and the aldosterone-induced increases in RSNA, HR, and MBP. Intracerebroventricular infusion of either the mineralocorticoid receptor antagonist RU-28318 or the AT(1)R antagonist losartan blocked aldosterone-induced phosphorylation of p44/42 MAPK and prevented the increases in RSNA, HR, and MBP. These data suggest that aldosterone-induced sympathetic excitation depends upon that AT(1)R-induced MAPK signaling in the brain. The short time course of this interaction suggests a nongenomic mechanism, perhaps via an aldosterone-induced transactivation of the AT(1)R as described in peripheral tissues.

Glucocorticoid Suppresses Steroidogenesis in Rat Progenitor Leydig Cells

Glucocorticoid (GC) inhibits testosterone production in adult Leydig cells by the glucocorticoid receptor (GR). However, whether GC affects the development of Leydig cells is unclear. The goal of the present study is to investigate the effects of GC on steroidogenesis of rat progenitor Leydig cells (PLCs) in vitro. Dexamethasone (DEX) inhibited androsterone (AO) production in PLCs. The GR antagonist RU38486 reversed the DEX-induced inhibition of AO, whereas the mineralocorticoid receptor antagonist RU28318 did not. RU38486 also reversed DEX-induced reductions in steady-state mRNA levels of steroidogenic acute regulatory protein (Star) and 3β-hydroxysteroid dehydrogenase 1 (Hsd3b1). Steroidogenic acute regulatory protein (StAR) protein expression and 3β-hydroxysteroid dehydrogenase (3βHSD) enzyme activity were affected similarly. These results show that GCs inhibit steroidogenesis of PLCs by suppression of StAR and 3βHSD via a GR-mediated mechanism.

Repeated restraint-induced modulation of long-term potentiation in the dentate gyrus of the mouse

We studied the potential for induction of long-term potentiation (LTP) in slices of the dentate gyrus of C57BL/6 mouse, prepared 24 h after 1, 3, 7, 14 or 21 successive daily neck restraint sessions lasting for 10 min. In slices from control animals and mice restrained only once, LTP (155 ± 2% of baseline) was evident for at least 2 h, but LTP was impaired after 3 sessions of neck restraint. This effect could be blocked by administration of the glucocorticoid receptor (GR) antagonist RU38486 before each restraint. One day after 3 restraint sessions the levels of plasma corticosterone, GR and mineralocorticoid receptor (MR) proteins remained elevated. After 7 neck restraint sessions the LTP level returned to normal; after 14 and 21 restraint sessions LTP was enhanced. LTP enhancement after 14 daily restraint sessions was not blocked by the GR antagonist RU38486 administered before each restraint; the MR antagonist RU28318 blocked LTP induction completely. After 14 daily restraint sessions the level of GR and MR proteins did not differ from the control and the plasma corticosterone level remained slightly elevated. These data indicate that repeated stress imposed by brief neck restraint bidirectionally modulates the potential for LTP in the dentate gyrus in a manner dependent on the number of daily restraint sessions.

Trilostane, FAD286, and the role of aldosterone in the central regulation of blood pressure: focus on “Role of central nervous system aldosterone synthase and mineralocorticoid receptors in salt-induced hypertension in Dahl salt-sensitive rats”

in a previous issue of this journal, Elise Gomez-Sanchez et al. ([4][1]) presented unequivocal evidence that intracerebroventricular (ICV) infusion of the 3β-hydroxy steroid dehydrogenase inhibitor trilostane prevented the development of hypertension in response to high-salt intake in Dahl salt-

Regulation of Alternative Splicing of Slo K+Channels in Adrenal and Pituitary during the Stress-Hyporesponsive Period of Rat Development

Stress triggers release of ACTH from the pituitary, glucocorticoids from the adrenal cortex, and epinephrine from the adrenal medulla. Although functions differ, these hormone systems interact in many ways. Previous evidence indicates that pituitary and steroid hormones regulate alternative splicing of the Slo gene at the stress axis-regulated exon (STREX), with functional implications for the calcium-activated K+ channels prominent in adrenal medullary and pituitary cells. Here we examine the role of corticosterone in Slo splicing regulation in pituitary and adrenal tissues during the stress-hyporesponsive period of early rat postnatal life. The sharp drop in plasma corticosterone (CORT) that defines this period offers a unique opportunity to test CORT's role in Slo splicing. We report that in both adrenal and pituitary tissues, the percentage of Slo transcripts having STREX declines and recovers in parallel with CORT. Moreover, addition of 500 nm CORT to cultures of anterior pituitary cells from 13-, 21-, and 30-d postnatal animals increased the percentage of Slo transcripts with STREX, whereas 20 microm CORT reduced STREX representation. Applied to adrenal chromaffin cells, 20 microm CORT decreased STREX inclusion, whereas neither 500 nm nor 2 microm had any effect. The mineralocorticoid receptor antagonist RU28318 abolished the effect of 500 nm CORT on splicing in pituitary cells, whereas the glucocorticoid receptor antagonist RU38486 blocked the effect of 20 microm CORT on adrenal chromaffin cells. These results support the hypothesis that the abrupt, transient drop in CORT during the stress-hyporesponsive period drives the transient decline in STREX splice variant representation in pituitary, but not adrenal.

Nongenomic effects of mineralocorticoid receptor activation in the cardiovascular system

Fifteen years ago Wehling and colleagues showed unequivocal rapid effects of aldosterone, neither mimicked by cortisol nor blocked by spironolactone, and postulated that these nongenomic effects are mediated via a membrane receptor distinct from the classical mineralocorticoid receptor (MR). Several recent studies have challenged this view. Alzamora et al. showed 11β-hydroxysteroid denydrogenase 1 and 2 (11βHSD1, 11βHSD2) expression in human vascular smooth muscle cells, and that aldosterone rapidly raises intracellular pH via sodium–hydrogen exchange; cortisol is without effect and spironolactone does not block the aldosterone response. When, however, 11βHSD activity is blocked by carbenoxolone, cortisol shows agonist effects indistinguishable from aldosterone; in addition, the effect of both aldosterone and cortisol is blocked by the open E-ring, water soluble MR antagonist RU28318. In rabbit cardiomyocytes, aldosterone increases intracellular [Na +] by activating Na +/K +/2Cl − cotransport, with secondary effects on Na +/K + pump activity. Pump current rises ∼10-fold within 15′, is unaffected by actinomycin D or the MR antagonist canrenone, and not elevated by cortisol. Pump current is, however, completely blocked by the open E-ring, water soluble MR antagonist K + canrenoate and stoichometrically by cortisol. PKCɛ agonist peptides (but not PKCα, PKCδ or scrambled PKCɛ peptides) mimic the effect of aldosterone, and PKCɛ antagonist peptides block the effect. Very recently, cortisol has been shown to mimic the effect of aldosterone when cardiomyocyte redox state is altered by the installation of oxidized glutathione (GSSG) via the pipet, paralleling the effect of carbenoxolone on vascular smooth cells and suggesting possible pathophysiologic roles for an always glucocorticoid occupied MR.

Nongenomic Cardiovascular Actions of Aldosterone: A Receptor for All Seasons?

Untilrecently,aldosteroneeffectshavebeendistinguished as acute nongenomic vs. more persistent genomic actions. Nongenomic actions are characterized by rapid onset and cannot be blocked by spironolactone nor mimicked by cortisol. On this basis, these rapid nongenomic effects of aldosterone were attributed to a membrane receptor, distinct from the classical mineralocorticoid receptor (MR), although such a receptor has not been characterized to date . Recently, however, Mano et al. (1) reported specific binding in plasma membranes of cardiac myocytes, with maximal levels of only 10.3 0.4 fmol/mg protein, and no correlation with functional data. The paper by Michea et al. (2) in the current issue of Endocrinology provides strong additional evidence that both the nongenomic and genomic actions of aldosterone, at least incardiovasculartissues,aremediatedviathesame,classical MR. The new mineralocorticoid antagonist, eplerenone, which binds specifically to the cytosolic receptor (3–6), blocks the rapid aldosterone effect on intracellular Ca 2 and pH as well as aldosterone-induced vasoconstriction. From other recent studies, there is a growing body of evidence that rapid nongenomic effects of aldosterone on vascular smooth muscle cells (7, 8) and cardiomyocytes (9, 10) are similarly mediated via classical MRs. A previous study from the Marusic laboratory (7) showed that human vascular smooth muscle cells express 11-HSD1 and 11-HSD2, and nanomolar concentrations of aldosterone acutely (5 min) increased intracellular pH in these cells. This effect was mediated by acute activation of the Na/H exchanger, as it was blocked by the amiloride derivative ethylisoprolpylamiloride but not spironolactone; in contrast, the water soluble, open E-ring MR antagonist RU28318 blocked the aldosterone-induced increase in pH. In keeping with other studies on the rapid effects of aldosterone, cortisol alone was without effect. When, however, the cortisol-metabolizing, reduced nicotinamide adenine dinucleotide-generating enzyme 11-HSD2 is blocked by addition of carbenoxolone, the glucocorticoid becomes active, mimicking the aldosterone action on intracellular pH.

Acute Exposure to a Novel Stressor Further Reduces the Habituated Corticosterone Response to Restraint in Rats

The present study sought to identify dishabituation of the hypothalamic-pituitary-adrenal (HPA) axis response to different psychological stressors. Young adult male Sprague Dawley rats were exposed to five, 1 h sessions of restraint stress on five consecutive days. On the sixth day, and 2 h before additional exposure to restraint, animals were subjected to 30 min of a small (27 cm square), elevated open field stressor (pedestal), which served as the dishabituating stimulus. We predicted HPA axis response dishabituation in chronically restrained rats exposed to the novel pedestal. Rats which underwent five days of restraint stress showed significantly blunted plasma corticosterone levels to restraint (habituation) as compared to restraint-naive rats. However, rats which underwent five sessions of restraint responded with an enhanced habituation response when confronted with restraint shortly after exposure to the novel pedestal. Instead of HPA axis response dishabituation, we observed enhanced habituation. Subsequent experiments determined that a 1.25mg/kg corticosterone injection could substitute for pedestal exposure to produce enhanced restraint habituation. Combined treatment with both the glucocorticoid receptor antagonist RU40555 (30 mg/kg) and the mineralocorticoid receptor antagonist RU28318 (50 mg/kg) blocked the expression of enhanced habituation after pedestal exposure. Thus, the delayed corticosterone negative feedback produced by novel stress led to enhanced expression of corticosterone response habituation.

Repeated blockade of mineralocorticoid receptors, but not of glucocorticoid receptors impairs food rewarded spatial learning.

Corticosteroids from the adrenal cortex influence a variety of behaviours including cognition, learning and memory. These hormones act via two intracellular receptors, the mineralo-corticoid receptor (MR) and the glucocorticoid receptor (GR). These two receptor types display a high concentration and distinct distribution in the hippocampus, a brain region which is directly involved in the regulation of spatial orientation and learning. In this study, repeated subcutaneous administration of the mineralocorticoid receptor antagonist RU28318 (1.0 mg/100 g body weight), the glucocorticoid receptor blocker RU38486 (2.5 mg/100 g body weight), or a combination of both antagonists were investigated for their effects on working--and reference memory in morning and afternoon trials during 8 subsequent days in food rewarded spatial learning in a hole board task. Each rat received one dose of either vehicle (2% ethanol in PEG 400), RU28318, RU38486 or the combination of both antagonists directly after the first trial on training days 1, 3, 5, and 7. The experiments demonstrated that repeated blockade of mineralocorticoid receptors impairs reference memory reflected in the morning--as well as in the afternoon trial, whereas blockade of glucocorticoid receptors has little effect on this type of cognitive behaviour. Furthermore, combined blockade of MRs and GRs resulted in a decrease, in both daily trials, in reference memory as well as working memory performance. These findings suggest that in this spatial learning paradigm, the impairment of working memory required blockade of both receptor types, while reference memory performance involves predominantly the mineralocorticoid receptors.

The resistance of the Wistar/Furth rat strain to steroid hypertension.

We have previously reported that the Wistar/Furth (W/Fu) rat strain is resistant to mineralocorticoid hypertension. In the current study, we have examined renal mRNA levels for mineralocorticoid receptor (MR), glucocorticoid receptor (GR), renin and Na+, K(+)-ATPase in response to treatment with mineralocorticoids. Uninephrectomized male Wistar (WI) and W/Fu rats were treated with aldosterone or deoxycorticosterone acetate (DOCA) and were given 1% NaCl to drink. Rats were sacrificed after 1, 3 or 7 days of treatment. Renal MR and ATPase mRNA levels were significantly reduced in aldosterone and DOCA-treated WI rats (e.g. MR was 30% on day 3 and ATPase was 50% of control on day 7 of aldosterone treatment). Unexpectedly, GR mRNA levels paralleled the changes in MR. In W/Fu rats the level of message was either unchanged or only moderately altered by this treatment. In vivo administration of the MR antagonist RU28318 or the GR antagonist RU38486 to WI rats for 4 days reduced renal mRNA levels for both subunits of ATPase. In the W/Fu rat, this treatment resulted in no change in the alpha subunit and an increase in the beta subunit of ATPase. In preliminary studies, we have determined that the W/Fu rat is also resistant to dexamethasone-induced hypertension. These studies suggest that altered MR- and GR-mediated mechanisms may contribute to the resistance of the W/Fu rat strain to steroid-induced hypertension.

11 alpha- and 11 beta-hydroxyprogesterone, potent inhibitors of 11 beta-hydroxysteroid dehydrogenase, possess hypertensinogenic activity in the rat.

The progesterone derivatives 11 alpha- and 11 beta-hydroxyprogesterone are potent inhibitors of 11 beta-hydroxysteroid dehydrogenase (isoforms 1 and 2) in vitro and can confer mineralocorticoid activity on corticosterone in the rat in vivo. 11 beta-Hydroxysteroid dehydrogenase metabolizes active glucocorticoids to their inactive 11-dehydro products and protects renal mineralocorticoid receptors from the high circulating levels of endogenous glucocorticoids. 11 beta-Hydroxysteroid dehydrogenase has been suggested to be important not only in the control of renal sodium retention but also of blood pressure. To assess the possible blood pressure-modulating effects of 11 alpha- and 11 beta-hydroxyprogesterone, we infused these substances into both intact and adrenalectomized Sprague-Dawley rats continuously for 14 days. Both 11 alpha- and 11 beta-hydroxyprogesterone caused a significant elevation in blood pressure within 3 days, an effect that persisted throughout the 14-day infusion. The hypertensive effects of 11 alpha-hydroxyprogesterone were abolished by adrenalectomy and significantly attenuated when 11 alpha-hydroxyprogesterone was infused together with the specific mineralocorticoid receptor antagonist RU28318. In an additional series of experiments, 11 alpha-hydroxyprogesterone significantly amplified the hypertensive effects of corticosterone in adrenalectomized spontaneously hypertensive rats but had no effects by itself in this experimental animal. These results demonstrate that both 11 alpha- and 11 beta-hydroxyprogesterone are potently hypertensinogenic in the rat and that this activity depends on an intact adrenal and at least in part on the activation of mineralocorticoid receptors. 11 beta-Hydroxyprogesterone, and similar endogenous progesterone metabolites that inhibit 11 beta-hydroxysteroid dehydrogenase, may be involved in the pathology of certain hypertensive states.

Determinants of cardiac fibrosis in experimental hypermineralocorticoid states.

Uninephrectomized rats maintained on 1.0% NaCl to drink and infused with aldosterone (0.75 microgram/h) for 8 wk have previously been shown to develop hypertension, cardiac hypertrophy, and cardiac fibrosis. In the present study we have shown that K+ supplementation (1.0% NaCl plus 0.4% KCl drinking solution) alters neither the interstitial nor the perivascular fibrotic response to mineralocorticoid treatment. Second, rats receiving 0.75 microgram/h 9 alpha-fluorocortisol, a mineralocorticoid and glucocorticoid agonist, respond with hypertension and cardiac fibrosis without cardiac hypertrophy. Finally, intracerebroventricular infusion of the mineralocorticoid receptor antagonist RU-28318 blocks blood pressure elevation, but not cardiac hypertrophy or fibrosis, when aldosterone is coinfused peripherally. We conclude that the myocardial fibrosis observed in response to chronic mineralocorticoid elevation and salt loading is a humorally mediated event independent of hypokalemia, hypertension, and cardiac hypertrophy. It remains to be determined whether the fibrosis observed in the presence of excess salt represents a direct (e.g., cardiac) effect of mineralocorticoid hormones or one mediated via a primary action on classical epithelial aldosterone target tissues (e.g., kidney).

Central effects of mineralocorticoid antagonist RU-28318 on blood pressure of DOCA-salt hypertensive rats.

The role of brain mineralocorticoid receptor (MR) sites in the pathogenesis of mineralocorticoid hypertension was studied after an intracerebroventricular injection of the MR antagonist RU-28318. Male Wistar rats received subcutaneously implanted deoxycorticosterone acetate (DOCA) pellets and were maintained on 0.9% saline as drinking solution. Under these conditions hypertension developed in approximately 5 wk as assessed in conscious rats by means of the tail-cuff technique. During the development of this hypertension (after 3 wk of DOCA-salt treatment) a single intracerebroventricular injection of the specific MR antagonist RU-28318 reduced systolic blood pressure (SBP) as measured with the tail-cuff method. A decrease in SBP was observed 2-24 h after this intracerebroventricular injection, with the lowest SBP values occurring at 8 h. In these animals (3 wk after DOCA implantation) continuous direct blood pressure recording via chronic cannulation revealed, on the day of the intracerebroventricular injection of RU-28318, a slight increase in arterial pressure during the light phase, followed by a decrease during the dark phase. In the established hypertensive rats (5 wk after DOCA RU-28318 on the arterial pressure or heart rate was detectable. It is concluded that central MR blockade during the development of the DOCA-salt hypertension reduces blood pressure within 24 h assessed with 1) the indirect method at certain time points after exposure to warming and stress and 2) the direct method during the dark phase of the diurnal cycle.

The effect of corticosterone on reactivity to spatial novelty is mediated by central mineralocorticosteroid receptors.

Corticosterone, secreted by the adrenal glands, binds to central mineralocorticoid receptors with high affinity and to glucocorticoid receptors with a tenfold lower affinity. In previous studies we have shown that the selective activation of either mineralocorticoid receptors or glucocorticoid receptors exerts distinctly different behavioural effects. In this study we examined in particular the mineralocorticoid receptor-mediated effect of corticosterone on the control of the behavioural response of male Wistar rats to spatial novelty. This analysis was based on our observation that in adrenal-intact rats the presence of an object in the centre of an open field alters the time spent and distance walked in the centre compared to the peripheral area, i.e. the pattern of reactive locomotor activity is changed. Using this paradigm we found that 1 day after removal of the adrenals the rats increased their behavioural reactivity towards the object. Treatment of adrenalectomized rats with a low dose of corticosterone (50 micrograms/kg s.c.) 1 h prior to testing restored the behavioural reactivity to the level of sham-operated, intact rats. Surprisingly, a high dose of corticosterone (1000 micrograms/kg s.c.) also increased the rat's reactivity towards the object. The same high dose of corticosterone given to adrenal-intact rats also increased behavioural reactivity. Pretreatment of these rats with an intracerebroventricular injection of the selective mineralocorticoid receptor antagonist RU28318 (100 ng/microliters) prevented the corticosterone-induced increase in behavioural reactivity, while the blockade of glucocorticoid receptors with the antagonist RU38486 (100 ng/microliters) was not effective. Administration of the mineralocorticoid receptor antagonist without corticosterone to adrenal-intact rats also increased behavioural reactivity, but this increase did not reach statistical significance.(ABSTRACT TRUNCATED AT 250 WORDS)

The apparent dependence of salt appetite in the pigeon on endogenous angiotensin II

Blockade of endogenous angiotensin II (ANG II) biosynthesis by intramuscular administration of the angiotensin converting enzyme inhibitor captopril (1 or 10 mg/kg b.wt.) completely suppressed salt appetite induced by sodium depletion in the pigeon. The effect was selective since captopril did not reduce deoxycorticosterone (DOCA)-induced salt appetite nor water drinking to ANG II and eledoisin. Blockade of brain ANG II receptors by pulse intracerebroventricular (pICV) injection of the ANG II receptor antagonist [Sarcosine 1, isoleucine 8] ANG II produced a marked, although partial, inhibition of salt appetite. The inhibition was quantitatively similar to the effectiveness of the ANG II receptor blockade, as measured by the suppression of drinking to pICV ANG II. Blockade of brain aldosterone (ALDO) receptors by pICV injections of the mineralocorticoid receptor antagonist RU-28318 did not significantly suppress depletion-induced appetite at doses that markedly reduced DOCA-induced salt appetite. These findings suggest that the pigeon might be completely dependent on ANG II for the expression of depletion-induced salt appetite. This is in contrast with what has been found in the rat, in which blockade of both ANG II and ALDO are necessary to suppress the appetite.