Fever evoked by macrophage inflammatory protein-1 (MIP-1) injected into preoptic or ventral septal area of rats depends on intermediary protein synthesis

Macrophage inflammatory protein-1 (MIP-1), a novel cytokine composed of α/β subunits, is released from macrophages during infection, MIP-1 injected intravenously in the rabbit or into the anterior hypothalamic, preoptic area (AH/POA) of the rat causes an intense fever, which is not blocked by prostaglandin synthesis inhibitors, ibuprofin or indomethacin, respectively. The purpose of this study was to determine the role of de novo protein synthesis on the fever evoked by MIP-1 applied to thermosensitive cells of the AH/POA. Guide cannulae were implanted bilaterally above the AH/POA or ventral septal area (VSA) and medially above the third cerebral ventricle in each of 11 male Sprague-Dawley rats. Following postoperative recovery, body temperature (T_b) was monitored by a colonic thermistor probe. The bilateral microinjection of MIP-1 in a dose of 14 pg per 0.5 μ1 into the AH/POA caused a biphasic elevation in T_b to 0.9 ± 0.2 °C within 3.0 h, and persisted for over 6.0 h. An identical injection of MIP-1 into the VSA increased T_b biphasically to 0.1 ± 0.1 °C within 1.0 h and to 0.8 ± 0.3 °C within 3.0 h. The infusion into the third ventricle of 80 μg/10 μ1 of the inhibitor of protein synthesis, anisomycin, either 10 or 30 min before the microinjection of MIP-1 into the AH/POA, attenuated significantly the rise in T_b for 1.0 to 3.0 h or 2.5 to 3.0 h, respectively. These results coincide with the earlier finding that anisomycin inhibits both endotoxin- and IL-1β-induced fevers. Further, the synthesis of a new protein factor may be required functionally for the initiation and maintenance of a fever whose mechanism of induction apparently is metabolically independent of the cyclooxygenase pathway.     

Cytokines and Thermoregulation: Interleukin-9 Injected in preoptic area fails to evoke fever in rats

A number of the members of the family of cytokines including IL-1, IL-2, IL-6, and IL-11 act directly in the brain to induce a febrile response in the rat and other species. The purpose of this study was to examine the effect of interleukin-9 (IL9) when this cytokine is applied directly to the thermosensitive and pyrogen reactive region of the anterior hypothalamic, preoptic area (AH/POA). In male Sprague-Dawley rats, guide cannulae for microinjection into the AH/POA were implanted stereotaxically, and radio transmitters for monitoring body temperature (T_b) were placed intraperitoneally. Following postoperative recovery, recombinant murine macrophage inflammatory protein (MIP)-1β was microinjected in the AH/POA of each rat in a dose of 28 pg/1μl to identify pyrogen reactive sites in the AH/POA. Then recombinant human IL-9 was suspended in pyrogen-free CSF vehicle and microinjected in the same sites in concentrations of 2.4, 24, and 240 U/μl. In contrast to the pyrexic action of MIP-1β, IL-9 failed to elicit a significant alteration in the T_b of the rats at any of the doses tested. IL-9 was also without effect on the intakes of either water or food. These results demonstrate that IL-9 applied to the region of the diencephalon in which other cytokines act to evoke fever may not play a direct role in the thermogenic component underlying the acute phase response. However, as demonstrated in several different cell systems, IL-9 may require a cofactor related to pyrogen for a febrile response to develop.     

Inhibition of interleukin-1β and prostaglandin E2 thermogenesis by glycyl-glutamine, a pro-opiomelanocortin-derived peptide

Interleukin-1β (IL-1β) and other cytokines produce fever by stimulating prostaglandin E₂ (PGE₂) synthesis in thermoregulatory regions of the preoptic area and anterior hypothalamus (POA/AH). Prostaglandin E₂ is thought to raise body temperature, at least in part, by stimulating β-endorphin release from pro-opiomelanocortin neurons that innervate the POA/AH. In this study, we investigated whether glycyl-glutamine (β-endorphin_30–31), an inhibitory dipeptide synthesized from β-endorphin post-translationally, inhibits IL-1β and PGE₂-induced hyperthermia. Hyperthermic sites were identified by microinjecting PGE₂ (3 fmol/1 μl) into the medial preoptic area (mPOA) of conscious, unrestrained rats. Interleukin-1β (1 U) injection into the same PGE₂ responsive thermogenic sites in the mPOA elicited a prolonged rise in colonic temperature (T_c) (+1.02±0.06°C) that persisted for at least 2 h. Glycyl-glutamine (3 nmol) co-injection into the mPOA inhibited IL-1β thermogenesis completely (T_c=−0.18±0.22°C). Glycyl-glutamine had no effect on body temperature when given alone to normothermic rats. Co-injection of individual amino acids, glycine and glutamine (3 nmol each amino acid), failed to influence IL-1β-induced thermogenesis, which indicates that Gly-Gln hydrolysis does not explain its inhibitory activity. Glycyl-glutamine (3 nmol) also prevented the rise in body temperature produced by PGE₂ (PGE₂=0.89±0.05°C; PGE₂ plus Gly-Gln=−0.16±0.14°C), consistent with evidence that PGE₂ mediates IL-1β-induced fever. These findings demonstrate that Gly-Gln inhibits the thermogenic response to endogenous pyrogens.     

Macrophage inflammatory protein-1: unique action on the hypothalamus to evoke fever

Macrophage inflammatory protein (MIP-1) administered systemically causes a fever not blocked by a prostaglandin (PGE) synthesis inhibitor. The purpose of this study was to examine the central mechanism of pyrexic action of this cytokine in the unrestrained rat. After guide cannulae for microinjection were implanted stereotaxically just above the anterior hypothalamic preoptic area (AH/POA), the body temperature of each rat was monitored by a colonic thermistor probe. Saline control vehicle or MIP-1 was microinjected into the AH/POA in one of eight concentrations ranging from 0.0028-9.0 ng per 0.5 mu 1 volume. MIP-1 induced a biphasic or monophasic fever of short latency characterized by an inverse dose-response curve. The potency of MIP-1 was in the femtomolar (10(-15)) range with the lowest dose of 0.028 ng producing a fever of over 2.0 degrees C with a latency of 15 min or less. To determine whether a PGE mediates MIP-1 fever, indomethacin was administered either intraperitoneally in a dose of 5.0 mg/kg or directly into the MIP-1 injection site in a dose of 0.5 microgram/0.5 mu 1, both injected 15 min before MIP-1. Pretreatment of the injection site in the AH/POA with indomethacin failed to prevent the febrile response evoked by MIP-1 injected at the same locus. Further, the dose of systemic indomethacin, which blocks PGE-induced fever in the rat, attenuated only partially the MIP-1 fever. The results demonstrate that MIP-1 is the most potent endopyrogen discovered thus far, and that its action is directly in the region of the hypothalamus which contains both thermosensitive and pyrogen-sensitive neurons. The local action of MIP-1 on cells of the AH/POA in evoking fever is unaffected by the PGE inhibitor which indicates, therefore, that a cellular mechanism operates in the hypothalamus to evoke fever independently of the central synthesis of a PGE.     

Sevoflurane neurotoxicity in neonatal rats is related to an increase in the GABAAR α1/GABAAR α2 ratio

Exposure of neonatal rat to sevoflurane leads to neurodegeneration and deficits of spatial learning and memory in adulthood. However, the underlying mechanisms remain unclear. The type A γ‐aminobutyric acid receptor (GABA_AR) is a target receptor for sevoflurane. The present study intends to investigate the changes in GABA_AR α1/α2 expression and its relationship with the neurotoxicity effect due to sevoflurane in neonatal rats. After a dose–response curve was constructed to determine minimum alveolar concentration (MAC) and safety was guaranteed in our 7‐day‐old neonatal rat pup mode, we conducted two studies among the following groups: (A) the control group; (B) the sham anesthesia group; and (C) the sevoflurane anesthesia group and all three groups were treated in the same way as the model. First, poly(ADP‐ribose) polymerase‐1 protein (PARP‐1) expression was determined in the different brain areas at 6 hr after anesthesia. Second, the expression of PARP‐1 and GABA_AR α1/GABA_AR α2 in the hippocampus area was tested by Western blotting at 6 hr, 24 hr, and 72 hr after anesthesia in all three groups. After 4 hr, with 0.8 MAC (2.1%) sevoflurane anesthesia, the PARP‐1 expression was significantly higher in the hippocampus than the other brain areas (p < .05). Compared with Groups A and B, the expression of PARP‐1 in the hippocampus of Group C significantly increased at 6 hr after sevoflurane exposure (216% ± 15%, p < .05), and the ratio of the α1/α2 subunit of GABA_AR surged at 6 hr (126% ± 6%), 24 hr (127% ± 8%), and 72 hr (183% ± 22%) after sevoflurane exposure in the hippocampus (p < .05). Our study showed that sevoflurane exposure of 0.8 MAC (2.1%)/4 hr was a suitable model for 7‐day‐old rats. And the exposure to sevoflurane could induce the apoptosis of neurons in the early stage, which may be related to the transmission from GABA_AR α2 to GABA_AR α1.     

Macrophage Inflammatory Protein-1β (MIP-1β) Produced Endogenously in Brain During E. coli Fever in Rats

Macrophage inflammatory protein-1 (MIP-1) evokes an intense fever, independent of a prostaglandin mechanism, and is now thought to play an important role in the defence response to bacterial pyrogens. The purpose of this study was 2-fold: (i) to determine whether the potent doublet of this cytokine, MIP-1β, is actually produced in the brain in response to a pyrogenic dose of a lipopolysaccharide of Escherichia coli and (ii) to determine the anatomical site of synthesis of this cytokine in the brain. Following the intense fever produced by intraperitoneal administration of lipopolysaccharide in the unrestrained rat, MIP-1β immunoreactivity was identified post mortem in two regions of the brain implicated in fever: the organum vasculosum laminae terminalis (OVLT) and the anterior hypothalamic, preoptic area (AH/POA). Microinjection of goat anti-mouse MIP-1β antibody (anti-MIP-1β) directly into the AH/POA markedly suppressed fever in rats in response to lipopolysaccharide. Further, anti-MIP-1β administered 180 min after the injection of lipopolysaccharide acted as an antipyretic and reversed the fever induced by the endotoxin. Anti-MIP-1β or control immunoglobulin G antibody microinjected into the hypothalamus immediately before the intraperitoneal injection of the control saline did not alter the temperature of the rats. Taken together, the present results demonstrate that MIP-1β is produced in the brain in response to a bacterial endotoxin. These observations, in the light of earlier data on fever induced by MIP-1β, further support the hypothesis that endogenously synthesized MIP-1β acts as an intermediary factor in the evocation of fever by acting on the thermosensitive cells of the brain.     

Antibodies to macrophage inflammatory protein-1β in preoptic area of rats fail to suppress PGE2 hyperthermia

This study determined whether macrophage inflammatory protein-1β (MIP-1β) plays a role in the hyperthermia caused by prostaglandin E₂ (PGE₂) given intracerebroventricularly (i.c.v.) in the rat. In these experiments, anti-murine MIP-1β antibody (anti-MIP-1β) was micro-injected in the anterior hypothalamic, preoptic area (AH/POA) just before i.c.v. PGE₂. The results showed that anti-MIP-1β failed to alter the PGE₂ hyperthermia. However, immunocytochemical studies revealed MIP-1β immunoreactivity detectable in both the organum vasculosum laminae terminalis (OVLT) and AH/POA in the febrile rat. These data thus demonstrate that MIP-1β is sequestered in diencephalic structures underlying thermoregulation even though it is not involved in PGE₂ hyperthermia. This dissociation supports the viewpoint that at least two distinct systems exist in the brain which underlie a febrile response: MIP-1β underlies one component whereas PGE₂ comprises the other.     

Blockade by interleukin-1 receptor antagonist of IL-1β-induced neuronal activity in guinea pig preoptic area slices

Interleukin-1α (IL-1α) and interleukin-1β (IL-1β) are thought to be endogenous pyrogens, i.e., to mediate fever production; warm-sensitive (W) and cold-sensitive (C) neurons in the preoptic area (POA) are presumed to be the ultimate targets of endogenous pyrogens. The recent purification of an IL-1 receptor antagonist (IL-1ra) has provided a means for verifying the presumptive action of IL-1 on these neurons. This study was undertaken, therefore, to investigate whether IL-1ra may block the IL-1α and IL-1β effects on the firing rates (FR) of W and C neurons in guinea pig POA slices. Human recombinant (hr) IL-1β (500 ng/ml) reduced the FR of 26 W neurons and increased those of 3 C neurons recorded; it had no effect on 8 thermally insensitive neurons. hrIL-1α (200–600 ng/ml) did not change the FR of any neuron. IL-1ra (0.01–0.5 mg/ml) had no effect by itself on the FR of all the neurons, but it blocked the hrIL-1β-induced FR changes of 24 of the 26 W and of all 3 C neurons when given before the cytokine. The lowest effective dose was 0.05 mg/ml. These results support the hypothesis, therefore, that POA thermosensitive neurons may be direct targets of IL-1β and that it may be an endogenous pyrogen acting on these units to induce fever production.     

Fever induced by macrophage inflammatory protein-1 (MIP-1) in rats: hypothalamic sites of action.

The purpose of this study was to clarify the central site of action as well as functional characteristics of the febrile response of the cytokine, macrophage inflammatory protein-1 (MIP-1). Guide cannulae for microinjection were implanted stereotaxically in the rat just above the pyrogen and thermosensitive area of the anterior hypothalamic, preoptic area (AH/POA). Following postoperative recovery, the body temperature of each rat (Tbo) was monitored during an experiment by a colonic thermistor probe at 0.5-1.0-h intervals. When MIP-1 was microinjected in a 0.5-microliter volume into the AH/POA in one of eight concentrations ranging from 0.0028 nanograms (ng) to 9.0 ng, an intense monophasic or biphasic fever was evoked. The MIP-1-induced increase in the Tbo of the rat was characterized by its short latency of 15 to 30 min and an inverse dose-response curve. Measures of mean latency and maximal rise in Tbo following MIP-1 confirmed the potency of this dose. Although the dose of 0.028 ng produced a fever of over 2.0 degrees C with a latency of only 15 min or less, the hyperthermic response became less intense as the dose of MIP-1 was increased. An anatomical mapping of sites of microinjection which reacted to MIP-1 in mediating fever revealed that the medial portion of the POA of the rat just rostral to the border of the AH was the region of maximum sensitivity to the cytokine.(ABSTRACT TRUNCATED AT 250 WORDS)     

Interleukin-1β (IL-1β)-induced modulation of the hypothalamic IL-1β system,tumor necrosis factor-α, and transforming growth factor-β1 mRNAs in obese (fa/fa) and lean (Fa/Fa) Zucker rats: Implications to IL-1β feedback systems and cytokine–cytokine interactions

Interleukin-1β (IL-1β) induces anorexia, fever, sleep changes, and neuroendocrine alterations when administered into the brain. Here, we investigated the regulation of the IL-1β system (ligand, receptors, receptor accessory protein, and receptor antagonist), tumor necrosis factor-α (TNF-α), transforming growth factor (TGF)-β1, and TGF-α mRNAs in the hypothalamus of obese (fa/fa) and lean (Fa/Fa) Zucker rats in response to the intracerebroventricular microinfusion of IL-1β (8.0 ng/24 hr for 72 hr, a dose that yields estimated pathophysiological concentrations in the cerebrospinal fluid). IL-1β increased IL-1β, IL-1 receptor types I and II (IL-1RI and IL-1RII), IL-1 receptor accessory protein soluble form (IL-1R AcP II), IL-1 receptor antagonist (IL-1Ra), TNF-α, and TGF-β1 mRNAs in the hypothalamus from obese and lean rats. IL-1β–induced IL-1β system and ligand (IL-1β, TNF-α, and TGF-β1) mRNA profiles were highly intercorrelated in the same samples. Levels of membrane-bound IL-1R AcP and TGF-α mRNAs did not change. Heat-inactivated IL-1β had no effect. The data suggest 1) the operation of an IL-1β feedback system (IL-1β/IL-1RI/IL-1R Acp II/IL-1RII/IL-1Ra) and 2) potential cytokine–cytokine interactions with positive (IL-1β ←→ TNF-α) and negative (TGF-β1 → IL-1β/TNF-α) feedback. Dysregulation of the IL-1β feedback system and the TGF-β1/IL-1β-TNF-α balance may have implications for neurological disorders associated with high levels of IL-1β in the brain. J. Neurosci. Res. 49:541–550, 1997. © 1997 Wiley-Liss, Inc.     

Anorexia and adipsia: dissociation from fever after MIP-1 injection in ventromedial hypothalamus and preoptic area of rats.

Certain cytokines such as tumor necrosis factor (TNF) and interleukin-1 (IL-1) act centrally to affect eating behavior and thermoregulation and may be involved in the physiological mechanisms leading to anorexia, adipsia and loss in body weight. The newly discovered macrophage inflammatory protein-1 (MIP-1) infused into the anterior hypothalamic, preoptic area (AH/POA) evokes an intense hyperthermia. The present experiments were designed to determine whether MIP-1 affects the feeding mechanism in the ventromedial hypothalamus (VMH) independently of the thermoregulatory mechanism in the AH/POA. For the microinjection of MIP-1, guide cannulae were implanted stereotaxically in the rat just above the VMH or AH/POA. Following postoperative recovery, each unrestrained rat was adapted to procedures whereby body temperature and intakes of food and water available ad lib were monitored at predetermined intervals. When an efficacious dose of 5.6 picograms (pg) MIP-1 was microinjected in a volume of 0.5 microliters into the VMH, the intake of food in the rat was reduced significantly in the short term and throughout the following 22 h. Within intervals of 30 min and 4.0 h following MIP-1, the amount of food consumed was 4.0 and 10 g, respectively, below that eaten by control rats given the saline solvent vehicle injected at the same site in the VMH. Over the entire test period, the intake of water was similarly significantly below that of the control rats. Whereas MIP-1 injected into the AH/POA evoked fever accompanied by a transient decline in feeding, the body temperature of the rats was unaffected by the cytokine injected in the VMH.(ABSTRACT TRUNCATED AT 250 WORDS)     

Neuregulin‐1β and neuregulin‐1α differentially affect the migration and invasion of malignant peripheral nerve sheath tumor cells

Malignant peripheral nerve sheath tumors (MPNSTs) are the most common malignancy associated with neurofibromatosis Type 1 (NF1). These Schwann cell lineage‐derived sarcomas aggressively invade adjacent nerve and soft tissue, frequently precluding surgical resection. Little is known regarding the mechanisms underlying this invasive behavior. We have shown that MPNSTs express neuregulin‐1 (NRG‐1) β isoforms, which promote Schwann cell migration during development, and NRG‐1α isoforms, whose effects on Schwann cells are poorly understood. Hypothesizing that NRG‐1β and/or NRG‐1α promote MPNST invasion, we found that NRG‐1β promoted MPNST migration in a substrate‐specific manner, markedly enhancing migration on laminin but not on collagen type I or fibronectin. The NRG‐1 receptors erbB3 and erbB4 were present in MPNST invadopodia (processes mediating invasion), partially colocalized with focal adhesion kinase and the laminin receptor β₁‐integrin and coimmunoprecipitated with β₁‐integrin. NRG‐1β stimulated human and murine MPNST cell migration and invasion in a concentration‐dependent manner in three‐dimensional migration assays, acting as a chemotactic factor. Both baseline and NRG‐1β‐induced migration were erbB‐dependent and required the action of MEK 1/2, SAPK/JNK, PI‐3 kinase, Src family kinases and ROCK‐I/II. In contrast, NRG‐1α had no effect on the migration and invasion of some MPNST lines and inhibited the migration of others. While NRG‐1β potently and persistently activated Erk 1/2, SAPK/JNK, Akt and Src family kinases, NRG‐1α did not activate Akt and activated these other kinases with kinetics distinct from those evident in NRG‐1β‐stimulated cells. These findings suggest that NRG‐1β enhances MPNST migration and that NRG‐1β and NRG‐1α differentially modulate this process. © 2009 Wiley‐Liss, Inc.     

α2, α2A, α2B/2C-Adrenoceptor subtype antagonists prevent lipopolysaccharide-induced fever response in rabbits

Exogenous pyrogens, e.g., bacterial lipopolysaccharides (LPS), are thought to stimulate macrophages to release endogenous pyrogens, e.g., TNFα, IL-1 β, and IL-6, which act in the hypothalamus to produce fever. We studied the effect of different α₁⁻ and α₂-adrenoceptor subtype antagonists, applied intraperitoneally, on the febrile response induced by LPS in rabbits. Evidence was obtained that prazosin, an α₁⁻ and α_2B/2C-adrenoceptor antagonist; WB-4101, an α₁⁻ and α_2A-adrenoceptor antagonist; CH-38083, a highly selective α₂-adrenoceptor antagonist (α₂: α₁ > 2000); BRL-44408, an α_2A-adrenoceptor antagonist; and ARC-239, an α_2B/2C⁻ and also α₁-adrenoceptor antagonist, blocked the increase of colonic temperature of the rabbit produced by 2 μg/kg LPS administered intravenously without being able in themselves to affect colonic temperature. In addition, prazosin, WB-4101 and CH-38083 antagonized the fall in skin temperature that occurred at the time when the colonic temperature was rising in control animals injected with LPS. All these results suggest that norepinephrine, through stimulation of both α₁⁻andα₂⁻ (α_2A⁻andα_2B/2C⁻) adrenoceptor subtypes, is involved in producing fever in response to bacterial LPS.     

A genome‐wide association study‐derived polygenic score for interleukin‐1β is associated with hippocampal volume in two samples

Accumulating research suggests that the pro‐inflammatory cytokine interleukin‐1β (IL‐1β) has a modulatory effect on the hippocampus, a brain structure important for learning and memory as well as linked with both psychiatric and neurodegenerative disorders. Here, we used an imaging genetics strategy to test an association between an IL‐1β polygenic score and hippocampal volume in two independent samples. Our polygenic score was derived using summary statistics from a recent genome‐wide association study of circulating cytokines that included IL‐1β (N = 3,309). In the first sample of 512 non‐Hispanic Caucasian university students (274 women, mean age 19.78 ± 1.24 years) from the Duke Neurogenetics Study, we identified a significant positive correlation between IL‐1β polygenic scores and hippocampal volume. This positive association was successfully replicated in a second sample of 7,960 white British volunteers (4,158 women, mean age 62.63 ± 7.45 years) from the UK Biobank. Our results lend further support in humans, to the link between IL‐1β and the structure of the hippocampus.     

Effects of Monoclonal Antibodies to Specific Epitopes of Rat Interleukin-1 Beta (IL-Iβ) on IL-1β-Induced ACTH, Corticosterone and IL-6 Responses in Rats

Recently, we developed a panel of monoclonal antibodies (MoAbs) to rat IL-1β and found that MoAbs binding to the aminoacid sequences 66–85 and 123–143 of mature rIL-1β inhibited the binding of rIL-1β to murine EL4 cells. Here we study whether MoAbs to these and other domains of IL-1 interfere with the biological effects of rIL-1β in adult male rats in vivo. Administration of rIL-1β (1 or 5 μg/kg i.v.) enhanced the plasma concentrations of ACTH, corticosterone (CORT) and of IL-6 in a time- (0.5–4 h) and dose-dependent manner. Because 2 h after 5μg/kg i.v., all three parameters were consistently elevated, this dose and time interval was used for further studies. Prior to injection, rIL-1β was incubated alone or in the presence of a MoAb (10 mg/kg) for 30 min at 37°C or at 4°C. Plasma ACTH, CORT and IL-6 responses to these mixtures are compared to those obtained after preincubation of rIL-1β with a non-IL-1 binding MoAb (PEN7). SILK 3, a MoAb that binds to the 66–85 domain of rIL-1β, reduced the ACTH and IL-6 responses by 48 and 45% respectively. In contrast, a MoAb to the 123–143 domain (SILK 5) and SILK 16, which binds to the 106–124 domain did not affect any of the IL-1 induced responses, whereas a MoAb directed to domain 78–97 of rIL-1β (SILK 20) enhanced the ACTH and CORT responses by 51 and 41% respectively, but not the IL-6 response. These observations lead us to conclude that the domain 66–85, harbours sequences that are important for receptor binding and for the biological actions of rIL-1β in the rat and that this biologically active domain is located at the ‘closed side’ rather than at the ‘open side’ of the IL-1β molecule where domains involved in receptor binding and biological activity are considered to be located.     

Activation of the hypothalamo-anterior pituitary corticotropin-releasing hormone,adrenocorticotropin hormone and β-endorphin systems during the estradiol 17β-induced plasma LH surge in the ovariectomized monkey

The present work describes time-dependent changes in the content of corticotropin-releasing hormone (CRH), adrenocorticotropin (ACTH), and β-endorphin (β -EP) in the hypothalamus (HT) and anterior pituitary (AP) and in the concentration of ACTH and β-EP in the plasma during the 17β estradiol (E2) benzoate (E₂B)-induced luteinizing hormone (LH) surge in ovariectomized cynomolgus monkeys. Monkeys were euthanized at 0, 30, 48, 72, and 96 hr post-E₂B. HT and AP were rapidly dissected, extracted in 2 N acetic acid containing 1 mM phenylmethane sulfonyl fluoride at 4°C, and centrifuged at 18,000g for 30 min. Peptide concentrations were measured in the supernatant by specific radioimmunoassays (RIAs). In the HT, there were significant (P < 0.05) decreases in ACTH and β -EP content by 30 hr post-E₂B and a significant (P < 0.05) decrease in HT CRH content 48 hr post-E₂B. Thereafter, CRH, ACTH, and β-EP content increased up to 72 hr post-E₂B. In the AP, there was an almost linear decrease in the CRH content through 48 hr post-E₂B followed by a marked 20-fold (P< 0.01) increase in the AP CRH content at 72 hr post-E₂B, which corresponds to the time of the descending arm of the LH surge. The patterns of ACTH and β-EP content were very similar in the AP, while that of CRH differed markedly. In contrast, in the HT CRH, ACTH, and β-EP profiles were very similar. Significant (P< 0.05) increases in circulating levels of ACTH, β-EP, and cortisol were evident at 30 hr (all 3 hormones), 48 hr (β-EP and cortisol), and 72 hr (cortisol) post-E₂B, which corresponds with the time of decreased hypothalamic content of CR1I, ACTH, and β-EP. These results suggest that there may be a marked activation of the hypothalamo-anterior pituitary-adrenal axis during the negative and positive feedback phases of the E₂B-induced LH surge in the ovariectomized monkey. © 1995 Wiley-Liss, Inc.     

Lack of effect of interleukins 1α and 1β, during in vitro perifusion,on anterior pituitary release of adrenocorticotropic hormone and β endorphin in the male rat

It has been demonstrated that interleukin 1 (IL1) injection provokes a great variety of biological effects, notably an activation of the corticotropic axis, increasing plasma adrenocorticotropic hormone (ACTH) and corticosterone. However, the primary site of action of IL1 is still controversial. In the present study, we first verified the in vivo capability of human interleukins 1α (hIL1α) and 1β (hIL1β) to release ACTH and β endorphin (β EP) in the normal male rat, before investigating, through an anterior pituitary (AP) perifusion system, the hIL1α and hIL1β effects on basal and corticotropin-releasing factor (CRF)-induced ACTH and β EP secretions. This system enabled the examination of a dynamic profile of hormones secretion, avoiding the possibility of feedback mechanisms, as is the case with the use of regular but very often longtime incubations. The results showed that in a perifusion system, with a short duration treatment (below 2 hr) compatible with the kinetics of action observed in vivo, basal and CRF-induced ACTH and β EP release were not modified in the presence of a broad range of concentrations (from 10^?12 to 10^?9 M) of hIL1α or hIL1β. Taken together, these results clearly show that in an in vitro situation close to physiological conditions, the primary site of action of hIL1α and hIL1β on ACTH and β EP release is not located at the AP level in the male rat. © 1993 Wiley-Liss, Inc.     

Relative efficacies of amyloid β peptide (Aβ) binding proteins in Aβ aggregation

The aggregation of amyloid β peptide (Aβ) into its fibrillar, cross β-pleated configuration is generally viewed as a critical event in the pathophysiology of Alzheimer's disease (AD). A diverse group of molecules, the Aβ binding proteins, has been evaluated for their effects on this process. However, most of these studies have used micromolar or greater reagent concentrations, and their different methods have not permitted quantitative comparisons of the efficacy of different Aβ binding proteins in augmenting or inhibiting aggregation. In the present work we have undertaken a coherent analysis using fluorimetry of thioflavin T-stained experimental solutions. The complement protein C1q, serum amyloid P, and transthyretin significantly enhanced the formation of precipitable, cross β-pleated aggregates in solutions of 800 nM Aβ1–42. Under these same experimental conditions, α₁-antichymotrypsin had no significant effect on the aggregation process, and both the E3 and E4 isoforms of apolipoprotein E were significant inhibitors. There was a non-significant trend toward the E3 isoform exhibiting greater inhibition than the E4 isoform. Of the aggregation-facilitating molecules, C1q was substantially and significantly the most potent. © 1996 Wiley-Liss, Inc.     

Enhanced aggregation and β structure of amyloid β peptide after coincubation with C1Q

Several lines of evidence now suggest that aggregation of soluble amyloid β peptide (Aβ) into a cross β sheet configuration may be an important factor in mediating potential neurotoxicity of Aβ. Synthetic Aβ has been shown to self aggregate in vitro. Here, we demonstrate that coincubation of freshly solubilized Aβ with C1q, a complement component known to bind Aβ in vitro and to colocalize with Aβ in vivo, results in as much as a 7-fold enhancement of Aβ aggregation, as well as a 2–4-fold enhancement of β structure within aggregates. The addition of C1q to preformed Aβ aggregates also results in significantly increased resistance to aggregate resolubilization. © 1994 Wiley-Liss, Inc.