Modulation of Interleukin-1 Receptors in the Brain-Endocrine-Immune Axis by Stress and Infection

We summarize data from some of our recent studies on in vitro and in vivo modulation of interleukin-1 (IL-1) receptors in the mouse brain-endocrine-immune axis by stress and infection. Ether-laparotomy stress in mice resulted in a selective increase in pituitary IL-I receptors and a significant decrease in pituitary receptors for corticotropin-releasing factor (CRF), a major regulator of the endocrine response to stress. Intraperitoneal injection of rat/human CRF mimicked the effects of stress and resulted in a dramatic increase in [¹²⁵I]IL-1α binding in the pituitary; [¹²⁵I]IL-1α binding in the hippocampus, spleen, and testis was unaffected by stress or CRF treatment. Glucocorticoid treatment with dexamethasone alone did not alter [¹²⁵I]IL-1α, binding but significantly inhibited CRF-induced upregulation of IL-1 receptors in the pituitary. The intracellular mechanism(s) involved in stress and CRF-induced upregulation of IL-1 receptors in the pituitary gland were examined by evaluating the effects of treatment of AtT-20 mouse pituitary corticotroph cells with a variety of neuroendocrine mediators of stress. CRF, forskolin, and isoproterenol (β₂ adrenergic receptor agonist) produced dose-dependent increases in cAMP production and [¹²⁵I]IL-1α binding. In contrast, somatostatin and dexamethasone significantly inhibited CRF-stimulated increase of cAMP production and [¹²⁵I]IL-1α binding, suggesting a primary role for cAMP in the regulation of pituitary IL-1 receptors. Next, we investigated the modulation of LL-1 β levels and IL-1 receptors following infection of mice with the endotoxin, lipopolysaccharide (LPS). Acute administration of low doses of endotoxin (30 μg LPS/mouse) dramatically increased IL-1 β levels and reciprocally decreased [¹²⁵I]IL-1α binding in peripheral tissues (pituitary, testis, liver, and spleen) but not in brain (hippocampus). This effect appeared to be dose related since higher doses of endotoxin (300 μg LPS/mouse) significantly decreased [¹²⁵I]IL-1α binding in both peripheral tissues and brain. Endotoxin induced modulation of the IL-1 system was also dependent on the treatment regimen since two low-dose LPS injections (at 0 and 12 h) increased IL-1 β concentrations and decreased [¹²⁵I]IL-1α binding in both central and peripheral tissues. These data provide further support for a role for IL-1 in coordinating brain-endocrine-immunoresponses to stress and infection.     

Differential transport of rat and human interleukin-1α across the blood–brain barrier and blood–testis barrier in rats

Human interleukin-1α is transported across the murine blood–brain barrier (BBB) and blood–testis barrier (BTB) by a saturable transport system. Differences in the biological activity and binding of human IL-1 in mouse and rat brain raise the possibility of species differences in the transport of IL-1 across the BBB and BTB. We measured the transport of recombinant human ¹²⁵I-IL-1α (I-huIL-1α) and rat ¹²⁵I-IL-1α (I-ratIL-1α) across the rat BBB and BTB after intravenous injection using a sensitive in vivo technique and film autoradiography. I-ratIL-1α was found to cross the rat BBB and rat BTB at rates comparable to those reported previously for murine IL-1α in mice. Passage across the BBB was inhibited by the addition of unlabeled rat IL-1α, demonstrating saturable transport. In contrast, I-huIL-1α entered the brain of the rat much more slowly, and its entry was not inhibited by the addition of unlabeled human IL-1α. These results show that the rat interleukin-1 transporter, unlike the murine transporter, does not transport human IL-1α. This difference highlights the importance of species specificity in IL-1α transport and may partly explain the different physiological responses to exogenous human IL-1α among rodent species.     

Elevation of plasma ACTH concentration in rabbits made febrile by systemic injection of bacterial endotoxin

This study was carried out to investigate the adrenocorticotrophic hormone (ACTH) response in rabbits made febrile by systemic injection of lipopolysaccharide (LPS,Salmonella typhosa endotoxin). Intravenous (i.v.) injection of LPS (0.1 μg/kg and 1.0 μg/kg) increased rectal temperature (biphasic fever) and the plasma concentration of ACTH (ACTH response) in a dose-related manner. These responses were suppressed by pretreatment with indomethacin (20 mg/kg, subcutaneously). Intracerebroventricular (i.c.v.) administration of indomethacin (400 μg) had no effect on the ACTH response to LPS, although it significantly suppressed febrile response. Small increases in plasma concentration of ACTH and significant fevers followed i.c.v. administration of prostaglandin E₂ (2 μg) or F_2α (2 μg). I.v. administration of corticotropin releasing factor (CRF) antagonist [α-helical CRF (9–41) (200 μg/kg)] partly suppressed the ACTH increased induced in plasma by i.v. LPS. These results suggest that prostagandins synthesized outside the blood-brain barrier play an important role in the ACTH response and that the mechanism for induction of the ACTH response is not exactly the same as that for the febrile response, although prostaglandins are involved in both responses.     

Effects of lipopolysaccharide and glucocorticoids on expression of interleukin-1β converting enzyme in the pituitary and brain of mice

The present study was carried out to determine whether those factors which regulate the expression of IL-1β in immune and non-immune tissues are also able to regulate the expression of ICE. In a first experiment, mice were injected with LPS (10 μg/mouse, ip) and killed before, 1, 3 or 6 h after the injection. Total RNAs were extracted from the spleen, pituitary and brain (hippocampus and hypothalamus) and submitted to RT-PCR to determine the levels of ICE mRNA as compared to β2microglobulin mRNA. ICE mRNAs were more abundant in the spleen and hippocampus than in the pituitary and hypothalamus but they were not significantly altered by LPS treatment. In a second experiment mice were submitted to adrenalectomy or a 15 min restraint stress and injected with saline or LPS (10 μg/mouse, sc). They were killed 1–2 h later and total RNA was extracted from the same tissues as in experiment 1. Adrenalectomized mice had significantly higher ICE mRNA levels whereas stressed mice had significantly lower ICE mRNA levels than their respective controls. These results are discussed with respect to the possible regulatory influence of glucocorticoids on the expression of ICE.     

ACTH- and α-MSH-induced grooming, stretching, yawning and penile erection in male rats: Site of action in the brain and role of melanocortin receptors

The effect of adrenocorticotropin (ACTH)(1-24) and α-melanocyte stimulating hormone (α-MSH) on grooming, stretching, yawning and penile erection was studied after injection into different brain areas. Both peptides induce the above responses when injected into the hypothalamic periventricular region of the third ventricle. This region includes the paraventricular nucleus, the dorsomedial nucleus, the ventromedial nucleus and the anterior hypothalamic area. The minimal effective dose of both peptides was 0.5 μg and the maximal effect was seen with 2 μg, the highest dose tested. Irrespective of the injection site, grooming started 5–7 min after injection of either peptide, while stretching, yawning and penile erection started only after 15–35 min and lasted for 90–120 min. In contrast both peptides were ineffective when injected into the preoptic area, the caudate nucleus or the CA1 field of the hippocampus. Grooming, stretching and yawning, but not penile erection, were prevented by cyclic[AcCys¹¹, D-Nal¹⁴, Cys¹⁸, AspNH₂²²]-β-MSH (11-22) (HS014), a selective melanocortin 4 receptor antagonist, injected into the same periventricular area 10 min before of ACTH(1-24) or α-MSH. The results show that ACTH(1-24) and α-MSH act in the hypothalamic periventricular region to induce the above responses and that grooming, stretching and yawning, but not penile erection, are mediated by melanocortin 4 receptors.     

In vivo Angiotensin II AT1 receptor blockade selectively inhibits LPS-induced innate immune response and ACTH release in rat pituitary gland

Systemic lipopolysaccharide (LPS) administration induces an innate immune response and stimulates the hypothalamic–pituitary–adrenal axis. We studied Angiotensin II AT₁ receptor participation in the LPS effects with focus on the pituitary gland. LPS (50 μg/kg, i.p.) enhanced, 3 h after administration, gene expression of pituitary CD14 and that of Angiotensin II AT_1A receptors in pituitary and hypothalamic paraventricular nucleus (PVN); stimulated ACTH and corticosterone release; decreased pituitary CRF₁ receptor mRNA and increased all plasma and pituitary pro-inflammatory factors studied.The AT₁ receptor blocker (ARB) candesartan (1 mg/kg/day, s.c. daily for 3 days before LPS) blocked pituitary and PVN AT₁ receptors, inhibited LPS-induced ACTH but not corticosterone secretion and decreased LPS-induced release of TNF-α, IL-1β and IL-6 to the circulation. The ARB reduced LPS-induced pituitary gene expression of IL-6, LIF, iNOS, COX-2 and IκB-α; and prevented LPS-induced increase of nNOS/eNOS activity. The ARB did not affect LPS-induced TNF-α and IL-1β gene expression, IL-6 or IL-1β protein content or LPS-induced decrease of CRF₁ receptors. When administered alone, the ARB increased basal plasma corticosterone levels and basal PGE₂ mRNA in pituitary.Our results demonstrate that the pituitary gland is a target for systemically administered LPS. AT₁ receptor activity is necessary for the complete pituitary response to LPS and is limited to specific pro-inflammatory pathways. There is a complementary and complex influence of the PVN and circulating cytokines on the initial pituitary response to LPS. Our findings support the proposal that ARBs may be considered for the treatment of inflammatory conditions.     

Somatostatin receptor-GTP binding regulatory protein-adenylyl cyclase system in hippocampal membranes of strychnine-treated rats

Wistar rats were injected with either a non-convulsive dose (37.5 μg/100 g body weight (b.wt.), intravenously (i.v.)) or a convulsive dose (50 or 80 μg/100 g b.wt., i.v.) of strychnine. Binding of¹²⁵I-Tyr¹¹-somatostatin (¹²⁵I-Tyr¹¹-SS) to its specific receptors was measured in hippocampal membranes 15 min after strychnine injection at these three doses. The non-convulsive dose of strychnine did not affect binding of SS in the hippocampus whereas both convulsive doses decreased the number of specific SS receptors without influencing their apparent affinity. Somatostatin-like immunoreactivity (SSLI), SS-modulated adenylel cyclase (AC) activity and the inhibitory guanine-nucleotide binding regulatory protein were also measured in rats treated with 80 μg/100 g b.wt. of strychnine. SSLI content remained stable. No significant differences were seen for the basal and forskolin (FK)-stimulated AC enzyme activities in the hippocampus of strychnine-treated rats when compared to the control group. The capacity of SS to inhibit basal and FK-stimulated AC activity in the hippocampus was significantly lower in the strychnine group than in the control group. The ability of the stable GTP analogue 5′-guanylylimidodiphosphate [Gpp(NH)p] to inhibit FK-stimulated AC activity was also decreased in hippocampal membranes from strychnine-treated rats. These results suggest that the attenuated inhibition of AC by SS in hippocampal membranes from strychnine-treated rats may be caused by decreases in both G_i activity and in the number of SS receptors. Alternatively, an uncoupling of SS receptors from G_i and/or a decrease in the level of functional G_i may result in both a decrease in apparentB_max for SS binding and in SS-mediated inhibition of AC. Since recent studies of other authors support the view that SS is predominantly an inhibitory transmitter in the hippocampus, it is possible that the decreased inhibition of AC activity by SS as well as the decrease in the number of SS receptors found in the hippocampus of strychnine-treated rats may be a mechanism involved in the development of increased seizure susceptibility.     

Estradiol, in the CNS, targets several physiologically relevant membrane-associated proteins

We will describe the identity and function of two unexpected estrogen binding proteins from rat brain cell membranes in search for the putative membrane estrogen receptor (mER). An E-6-BSA column retained a distinctive 37-kDa protein that showed 100% homology with glyceraldehyde-3-phosphate dehydrogenase (GAPDH). A P-3-BSA column also retained the same protein but with less affinity. E-6-BSA bound to GAPDH with an IC₅₀ of 50 nM, whereas the IC₅₀ for P-3-BSA was about 500 nM. A dose of 10 nM 17β-estradiol stimulated the catalysis of GAPDH, whereas progesterone at 100 nM inhibited it. Other steroids were ineffective. We examined if GAPDH activity would change during the rat estrous cycle, and what would be the effect of ovariectomy and estrogen treatment. The hippocampus and cerebellum were collected and GAPDH catalysis in both cytosolic and plasmalemmal-microsomal fractions was tested. The highest activity was found in Proestrus morning and the lowest in Estrus in both fractions. After ovariectomy (3 weeks) the hippocampus membrane fraction showed significantly reduced activity compared to that of Diestrus. An injection of estradiol in ovariectomized rats (10 μg/rat, s.c.) increased GAPDH activity in the hippocampus membrane fractions close to 60% from that of ovariectomized oil-treated controls 24 h after treatment maintaining similar levels by 48 h. No changes were detected in the preparations from the cerebellum of the same rats. The other protein retained by E-BSA columns was a 55-kDa protein identified as β-tubulin. Two other proteins were also co-purified from the rat hippocampus: a 37-kDa (GAPDH) and a 45-kDa (actin). A purified brain tubulin (Cytoskeleton) was also retained with high affinity by the E-6-BSA, but with less affinity by an E-17-BSA column and not retained by either BSA, P-3-BSA or C-21-BSA columns. E-6-[¹²⁵I]BSA bound with high affinity to tubulin (1 μg) and 17β-estradiol completely displaced the binding at 10⁻⁷ M. 17α-Estradiol was ineffective and neither progesterone, corticosterone, DES nor 2-methoxyestradiol (2-ME) was able to displace the ligand. The T-3-[¹²⁵I]BSA also bound to tubulin. But it seems to interact with another binding site, because colchicine at 10⁻⁵ M completely eliminated the binding of T-3-[¹²⁵ I]BSA to tubulin but did not displace the E-6-BSA site. Taxol competed off both ligands but only by 50%. None of the two ligands bound actin. These novel findings add new information to be considered in the intracellular actions of estradiol, particularly in the remodeling and functions of the cytoskeleton.     

Opioid and cannabinoid receptor-mediated regulation of the increase in adrenocorticotropin hormone and corticosterone plasma concentrations induced by central administration of Δ-tetrahydrocannabinol in rats

The purpose of this study was to investigate the cannabinoid and opioid mediated regulation on the effects of central Δ⁹-tetrahydrocannabinol (Δ⁹-THC) administration on hypothalamus–pituitary–adrenal (HPA) axis activity in the male rat. Intracerebroventricular (i.c.v.) administration of Δ⁹-THC (25, 50, 100 μg/rat) markedly increased plasma adrenocorticotropin hormone (ACTH) and corticosterone concentrations. Time course effect studies revealed that both hormones secretion peaked at 60 min after Δ⁹-THC i.c.v. administration (50 μg/rat), decreased gradually and returned to baseline levels by 480 min. The i.c.v. administration of the specific cannabinoid receptor antagonist SR-141716A (3 μg/rat) significantly attenuated the increase of both hormones secretion induced by Δ⁹-THC (50 μg/rat). Nevertheless, higher doses (12.5 and 50 μg/rat) of this compound increased both ACTH and corticosterone plasma concentrations. Subcutaneous (s.c.) administration with the opiate receptor antagonist naloxone (0.3 mg/kg) was without effect but significantly diminished the increase of both hormones secretion induced by Δ⁹-THC (50 μg/rat). Taken together, these results indicate that opiate and cannabinoid receptors are involved in the activation of the HPA axis induced by Δ⁹-THC. Furthermore, the increase of ACTH and corticosterone secretion after the administration of higher doses of SR-141716A than those required to block such activation, suggests that endogenous cannabinoids are tonically inhibiting the release of both hormones or that this agonist-like activity may be part of an uncharacterized action of this compound not mediated by cannabinoid receptors.     

Social stress inhibits the nitric oxide effect on the corticotropin-releasing hormone- but not vasopressin-induced pituitary–adrenocortical responsiveness

Putative involvement of endogenous nitric oxide (NO) in the corticotropin-releasing hormone (CRH, 1 μg/kg i.p.)- and vasopressin (AVP, 5 μg/kg i.p.)-induced ACTH and corticosterone secretion was investigated in both non-stressed and crowded rats. The NO synthase blocker Nω-nitro- -arginine ( -NNA, 2 mg/kg i.p.) significantly augmented the AVP-induced ACTH and corticosterone secretion in control and stressed rats, but it increased the CRH-induced ACTH response only in control rats. Crowding stress did not affect the -NNA evoked increase in AVP-induced hormone responses, but it abolished the CRH-induced ACTH response.     

α-Bungarotoxin binding sites in rat hippocampal and cortical cultures: initial characterisation, colocalisation with α7 subunits and up-regulation by chronic nicotine treatment

High density neuronal cultures from rat E18 hippocampus and cortex have been characterised with respect to cholinergic binding sites. No specific binding of [³H]nicotine or [³H]cyttine to live cells in situ was detected, although the limit for detection was estimated to be 30 fmol/mg protein. Muscarinic binding sites labelled with [³H]QNB were present at a density of 0.75 pmol/mg protein. [¹²⁵I]α-Bungarotoxin (αBgt) bound to hippocampal cultures with a B_max of 128 fmol/mg protein and a K_d of 0.6 nM; cortical cultures expressed five times fewer [¹²⁵I]α-Bgt binding sites. Fluorescence cytochemistry with rhodamine-α-Bgt indicated that 95% of hippocampal neurons were labelled, compared with only 36% of cortical neurons. Average densities of 4 × 10⁴ and 2 × 10⁴ binding sites/cell were calculated for hippocampal and cortical cultures, respectively. Double labelling experiments with mAb307 (which recognises the rat α7 nicotinic receptor subunit) and rhodamine-α-Bgt gave coincident labelling patterns, supporting the correlation between the α7 subunit and Bgt-sensitive neuronal nicotinic receptor. Treatment of hippocampal cultures with 10 μM nicotine for 14 days elicited a 40% increase in the numbers of [¹²⁵I]α-Bgt binding sites, mimicking the up-regulation observed in vivo studies. Primary cultures offer a useful in in vitro system for investigating the expression and regulation of brain α-Bgt-sensitive receptors.     

Distribution of an α-bungarotoxin-binding cholinergic nicotinic receptor in rat brain

Cholinergic nicotinic receptors in rat brain were demonstrated by the use of the potent nicotinic antagonist [¹²⁵I]α-bungarotoxin ([¹²⁵I]α-Btx). Biochemical studies on binding of [¹²⁵I]α-Btx to rat hippocampal homogenates revealed saturable binding sites which are protected by nicotine, d-tubocurarine and acetylcholine but not by atropine or oxotremorine. The hippocampus and hypothalamus displayed relatively high [¹²⁵I]α-Btx specific binding whereas the cerebellum was devoid of specific binding. Other regions displayed intermediate binding levels. Analysis of the regional distribution of [¹²⁵I]α-Btx binding by autoradiography of frontal brain sections revealed high labeling in the hippocampus, hypothalamic supraoptic, suprachiasmatic and periventricular nuclei, ventral lateral geniculate and the mesencephalic dorsal tegmental nucleus. It is suggested that the limbic forebrain and midbrain structures as well as sensory nuclei are the main nicotinic cholinoceptive structures in the brain.     

Pharmacological characterisation of the histamine H3 receptor in the rat hippocampus

The purpose of this report was to pharmacologically characterise the histamine H₃ in the rat hippocampus using radioligand binding studies with the H₃ receptor antagonist [¹²⁵I]iodophenpropit and the H₃ receptor mediated inhibition of [³H]noradrenaline release. A dissociation constant of 0.33 nM and a maximal number of binding sites of 125 fmol/mg protein were found for [¹²⁵I]iodophenpropit. Competition studies showed stereoselectivity for the (R) and (S) enantiomers of α-methylhistamine and 10 μM of GTP_γS shifted the curve of (R)-α-methylhistamine rightwards. Up to 1 μM, (R)-α-methylhistamine displaced only 30% whereas the tested H₃-antagonists displaced 50–60% of the total [¹²⁵I]iodophenpropit bound. This indicates the presence of an additional non-H₃ receptor binding site(s) for [¹²⁵I]iodophenpropit in the rat hippocampus. This secondary site shows low affinity for H₃ agonists, but high affinity for the tested H₃ antagonists. Electrically evoked [³H]acetylcholine release was shown in slices of rat hippocampus. No H₃ receptor modulation of [³H]acetylcholine release from hippocampal slices was detectable. However, H₃ receptor activation inhibited 42% of the electrically-evoked [³H]noradrenaline release in rat hippocampal slices. The inhibition of [³H]noradrenaline release was effectively antagonized by the H₃ antagonists thioperamide and burimamide. We describe the pharmacological identification of the histamine H₃ receptor in the rat hippocampus and its similarities and differences from the cortical H₃ receptor. These studies enable us to investigate changes in density and functionality of the hippocampal H₃ receptor under (patho)physiological conditions.     

Hippocampal cytosol binding capacity of corticosterone: no depletion with nuclear loading

The recurrence every 24 h of glucocorticoid elevation led us to investigate the temporal relationship between glucocorticoid receptor occupation in brain cell nuclei and the availability of cytosol sites. Adrenalectomized rats were injected i.v. with [³H]corticosterone in doses ranging from 15 to 106 nmol/kg. Peak nuclear binding occured 1–2 h after [³H]corticosterone injection and was preceded by a peak of cytosol receptor labeling at 15–30 min. Yet 1 h after 55 or 105 nmol/kg [³H]-corticosterone, no depletion in the total in vitro binding capacity of the cytosol could be detected even though estimated depletion should have been ≈30% had it occured. Injection of 80 nmol/kg of corticosterone per rat plus 40 nmol/kg dexamethasone also failed to reduce total cytosol binding capacity, even though estimated depletion should have been ≈40%. No change in binding affinity of cytosol sites was observed in the injected animals compared to uninjected controls.The in vivo nuclear binding capacity of hippocampus for [³H]corticosterone (fmol/hippocampus) is about 40% of the cytosol binding capacity measured in vitro. Moreover, no more than 40% of total cytosol sites are occupied in vivo as a result of [³H]corticosterone injections which occupy nuclear sites to 80% of estimated capacity. Yet, even with the larger in vitro cytosol binding capacity, a depletion approaching 40% of cytosol binding sites would have been seen, had it occurred as a result of nuclear translocation. The apparent lack of depletion of cytosol receptors is supported by experiments which showed that two injections of [³H]corticosterone 2 h apart fail to fatigue the nuclear uptake mechanism. The present results suggest (1) that in the hippocampus an excess of extranuclear glucocorticoid binding proteins exists, and (2) that the availability of functional cytosol receptors may be regulated to maintain a relatively constant cellular level.     

Differential modulation of angiotensin II and hypertonic saline-induced drinking by opioid receptor subtype antagonists in rats

Opioid modulation of ingestion includes general opioid antagonism of different forms of water intake, μ₂ receptor modulation of deprivation-induced water intake and δ₂ receptor modulation of saccharin intake. Water intake is stimulated by both central administration of angiotensin II (ANG II) and peripheral administration of a hypertonic saline solution; both responses are reduced by general opioid antagonists. The present study examined whether specific opioid receptor subtype antagonists would selectively alter each form of water intake in rats. Whereas systemic naltrexone (0.1–2.5 mg/kg, s.c.) reduced water intake induced by either peripheral ANGII (500 μg/kg, s.c.) or hypeptonic saline (3 ml/kg, 10%), intracerebroventricular (i.c.v.) naltrexone (1–50 μg) only inhibited central ANGII (20 ng)-induced hyperdipsia. Both forms of drinking were significantly and dose-dependently inhibited by the selective κ antagonist, nor-binaltorphamine (Nor-BNI, 1–20 μg). Whereas both forms of drinking were transiently reduced by the μ-selective antagonist, β-funaltrexamine (β-FNA, 1–20 μg), the μ₁ antagonist, naloxonazine (40 μg) stimulated drinking following hypertonic saline. The δ₁ antagonist, [d-Ala², Leu⁵, Cys⁶]-enkephalin (DALCE, 1–40 μg) significantly reduced drinking following ANGII, but not following hypertonic saline; the δ antagonist, naltrindole failed to exert significant effects. These data indicate that whereas κ opioid binding sites modulate hyperdipsia following hypertonic saline, μ₂, δ₁ and κ opioid binding sites modulate hyperdipsia following ANGII. The μ₁ opioid binding site may normally act to inhibit drinking following hypertonic saline.     

The role of interleukin-6 in the activation of the hypothalamo-pituitary-adrenocortical axis and brain indoleamines by endotoxin and interleukin-1β

Interleukin-6 (IL-6) is one of several cytokines that can stimulate the hypothalamo-pituitary-adrenocortical (HPA) axis. Because IL-6 is produced in response to the administration of endotoxin (LPS) and interleukin-1 (IL-1), it is possible that IL-6 contributes to the neuroendocrine and neurochemical changes induced by them. In this study, intraperitoneal (i.p.) injection of LPS elevated plasma concentrations of IL-6 while activating the HPA axis in a dose-dependent manner. Both responses reached a peak at around 2–3 h. Mouse IL-1β administration (100 ng, i.p.) induced large increases in plasma corticosterone and a substantial, but short-lived increase in plasma IL-6 with a peak at 2 h. Pretreatment of mice intraperitoneally with a monoclonal antibody to mouse IL-6 significantly attenuated the plasma ACTH and corticosterone responses to LPS at 3 h, but not at 1 h. Anti-IL-6 treatment also attenuated the LPS-induced increases of tryptophan and the serotonin catabolite, 5-hydroxyindoleacetic acid (5-HIAA), but not that of the norepinephrine catabolite, 3-methoxy,4-hydroxyphenylethyleneglycol (MHPG). Pretreatment of mice with anti-IL-6 significantly attenuated the IL-1-induced increases of plasma ACTH and corticosterone at 2 h, but not at 4 h. The IL-1-induced increases of MHPG, tryptophan and 5-HIAA in hypothalamus and brain stem were not significantly altered. These results suggest that IL-6 contributes to the later phases of the LPS- and IL-1-induced stimulations of the HPA axis and to the indoleaminergic responses to LPS, but not to IL-1.     

Neuropeptide K enhances glucocorticoid release by acting directly on the rat adrenal gland: the possible involvement of zona medullaris

Neuropeptide K (NPK), a member of the kassinin-like tachykinin family, is contained in the rat hypothalamus and is known to stimulate pituitary ACTH release. The intraperitoneal bolus administration of NPK dose-dependently enhanced corticosterone blood level not only in intact rats, but also in hypophysectomized/ACTH replaced animals. NPK did not affect corticosterone secretion of dispersed rat adrenocortical cells: however, it concentration-dependently raised basal corticosterone production by decapsulated adrenal quarters (including both cortical and medullary tissues). Minimal and maximal effective concentrations were 10⁻⁹ and 10⁻⁸ M, respectively. 10⁻⁸ M NPK potentiated corticosterone response of adrenal quarters elicited by 10⁻¹² M ACTH, but not that evoked by higher concentrations of ACTH. The direct corticosterone secretagogue effect of 10⁻⁸ M NPK is annulled by 10⁻⁶ M α-helical-CRH or corticotropin-inhibiting peptide, competitive inhibitors of CRH and ACTH, respectively. In light of these findings, the hypothesis is advanced that NPK exerts a direct stimulatory action on adrenocortical secretion and that the mechanism underlying this effect of NPK may involve the activation of the intra-medullary CRH/ACTH system.     

Corticotropin-releasing factor antagonist blocks microwave-induced decreases in high-affinity choline uptake in the rat brain

Acute (45-min) irradiation with pulsed low-level microwaves (2450-MHz, 2 μsec pulses at 500 pps, average power density of 1 mW/cm², whole-body average specific absorption rate of 0.6 W/kg) decreased sodium-dependent high-affinity choline uptake (HACU) activity in the frontal cortex and hippocampus of the rat. These effects were blocked by pretreating the animals before exposure with intracerebroventricular injection of the specific corticotropin-releasing factor (CRF) receptor antagonist, α-helical-CRF_9–41 (25 μg). Similar injection of the antagonist had no significant effect on HACU in the brain of the sham-exposed rats. These data suggest that low-level microwave irradiation activates CRF in the brain, which in turn causes the changes in central HACU.     

Uptake and retention of androgen in neurons of the brain of the golden hamster

The distribution of cells capable of concentrating androgen was studied in the male hamster after injection of 5α-dihydro-[1,2,4,5,6,7,(n)-³H]testosterone([³H]DHT) using the technique of thaw-mount autoradiography. Castrated adult male hamsters were injected with 0.2 μg/100 g body weight of [³H]DHT (107 Ci/mmol) and killed 1.5 h later. Brains were rapidly removed and processed for autoradiography. Localization of radioactivity in high concentrations occurred chiefly in limbic forebrain structures and hypothalamic nuclei associated with the control of reproductive function including the following areas: the septal-preoptic region, the amygdala, and the anterior, ventromedial and arcuate nuclei of the hypothalamus. In addition, labeled cells in lesser concentrations were found in the lateral preoptic area, lateral hypothalamus, hippocampus, mesencephalon and various cortical regions. Treatment with 100-fold excess of testosterone, but not estradiol or diethylstilbestrol, inhibited nuclear localization. These studies provide information on the precise anatomical localization of androgen concentrating cells in the hamster brain and demonstrate the similarity of distribution of anrdrogen binding in the rat, mouse and hamster.