Serotonin-type 3 receptors mediate intestinal lipid-induced satiation and Fos-like immunoreactivity in the dorsal hindbrain

Several gastrointestinal stimuli, including some intestinal nutrients, have been shown to exert their satiating effect via activation of serotonin type-3 (5-HT(3)) receptors. The presence of lipids in the small intestine potently suppresses food intake; however, whether 5-HT(3) receptors play a role in this response has not been directly examined. Therefore, using the selective 5-HT(3) receptor antagonist ondansetron, we tested the hypothesis that duodenal infusion of lipid suppresses intake of both sucrose solution and chow through 5-HT(3) receptor activation. Rats duodenally infused with 72 and 130 mM Intralipid suppressed 1-h 15% sucrose intake by 33 and 67%, respectively. Suppression of sucrose intake by 72 mM Intralipid was significantly attenuated by ondansetron at all doses tested (0.5, 1.0, 2.0, and 5.0 mg/kg ip), whereas the lowest effective dose of ondansetron to attenuate suppression of intake by 130 mM Intralipid was 1.0 mg/kg. Furthermore, infusion of 130 mM Intralipid suppressed 1- and 4-h chow intake by 35 and 20%, respectively. Ondansetron administered as low as 0.5 mg/kg significantly attenuated 1-h Intralipid-induced suppression of chow intake and completely reversed the suppression by 4 h. Administration of ondansetron alone did not alter sucrose or chow intake compared with vehicle injection at any time. Finally, to test whether Intralipid-induced neuronal activation of the dorsal vagal complex is mediated by 5-HT(3) receptors, Fos-like immunoreactivity (Fos-LI) was quantified in ondansetron-pretreated rats following intestinal lipid infusion. Ondansetron (1 mg/kg) significantly attenuated duodenal intralipid-induced Fos-LI in the dorsal hindbrain. These data support the hypothesis that 5-HT(3) receptors mediate both satiation, as well as hindbrain neuronal responses evoked by intestinal lipids.     

Serotonin 5-HT(7) receptors mediate relaxation of porcine pial veins

Isolated porcine pial veins in the presence of active muscle tone have been shown to exhibit rhythmic contractions (RC) that are inhibited by serotonin (5-HT) in a concentration-dependent manner. The 5-HT inhibition of RC is mediated by an as yet unidentified 5-HT receptor subtype located on the vascular smooth muscle. 5-carboxamidotryptamine, which is a potent but nonselective agonist at 5-HT(7) receptors, has been shown to be the most potent inhibitor of RC in porcine pial veins. Therefore, the present study was designed to determine if the 5-HT-mediated inhibition of RC in pial veins is mediated by 5-HT(7) receptors and if 5-HT(7) receptor mRNA is expressed in endothelium-denuded pial veins; the study was done with the use of an in vitro tissue bath and RT-PCR techniques. Our findings indicated that 5-HT inhibition of RC in porcine pial veins was prevented by 5-HT(7)-receptor antagonists (clozapine, pimozide, and LY-215840) in a concentration-dependent manner. Furthermore, a strong PCR signal for the 5-HT(7) receptor was consistently detected in endothelium-denuded pial veins. Sequence analysis of the amplified products confirmed their high degree of homology with the porcine and/or human 5-HT(7)-receptor gene. Taken together, these data suggest that the 5-HT-induced inhibition of RC in porcine pial veins is at least in part mediated by 5-HT(7) receptors located on the venous smooth muscle.     

Serotonin augments gut pacemaker activity via 5-HT3 receptors

Serotonin (5-hydroxytryptamine: 5-HT) affects numerous functions in the gut, such as secretion, muscle contraction, and enteric nervous activity, and therefore to clarify details of 5-HT's actions leads to good therapeutic strategies for gut functional disorders. The role of interstitial cells of Cajal (ICC), as pacemaker cells, has been recognised relatively recently. We thus investigated 5-HT actions on ICC pacemaker activity. Muscle preparations with myenteric plexus were isolated from the murine ileum. Spatio-temporal measurements of intracellular Ca(2+) and electric activities in ICC were performed by employing fluorescent Ca(2+) imaging and microelectrode array (MEA) systems, respectively. Dihydropyridine (DHP) Ca(2+) antagonists and tetrodotoxin (TTX) were applied to suppress smooth muscle and nerve activities, respectively. 5-HT significantly enhanced spontaneous Ca(2+) oscillations that are considered to underlie electric pacemaker activity in ICC. LY-278584, a 5-HT(3) receptor antagonist suppressed spontaneous Ca(2+) activity in ICC, while 2-methylserotonin (2-Me-5-HT), a 5-HT(3) receptor agonist, restored it. GR113808, a selective antagonist for 5-HT(4), and O-methyl-5-HT (O-Me-5-HT), a non-selective 5-HT receptor agonist lacking affinity for 5-HT(3) receptors, had little effect on ICC Ca(2+) activity. In MEA measurements of ICC electric activity, 5-HT and 2-Me-5-HT caused excitatory effects. RT-PCR and immunostaining confirmed expression of 5-HT(3) receptors in ICC. The results indicate that 5-HT augments ICC pacemaker activity via 5-HT(3) receptors. ICC appear to be a promising target for treatment of functional motility disorders of the gut, for example, irritable bowel syndrome.     

Serotonin 5-HT3 receptors in the central nervous system

The 5-HT₃ receptor is a ligand-gated ion channel activated by serotonin (5-HT). Although originally identified in the peripheral nervous system, the 5-HT₃ receptor is also ubiquitously expressed in the central nervous system. Sites of expression include several brain stem nuclei and higher cortical areas such as the amygdala, hippocampus, and cortex. On the subcellular level, both presynaptic and postsynaptic 5-HT₃ receptors can be found. Presynaptic 5-HT₃ receptors are involved in mediating or modulating neurotransmitter release. Postsynaptic 5-HT₃ receptors are preferentially expressed on interneurons. In view of this specific expression pattern and of the well-established role of 5-HT as a neurotransmitter shaping development, we speculate that 5-HT₃ receptors play a role in the formation and function of cortical circuits.     

Epidermal growth factor and betacellulin mediate signal transduction through co-expressed ErbB2 and ErbB3 receptors.

Interleukin-3 (IL-3)-dependent murine 32D cells do not detectably express epidermal growth factor receptors (EGFRs) and do not proliferate in response to EGF, heregulin (HRG) or other known EGF-like ligands. Here, we report that EGF specifically binds to and can be crosslinked to 32D transfectants co-expressing ErbB2 and ErbB3 (32D.E2/E3), but not to transfectants expressing either ErbB2 or ErbB3 individually. [125I]EGF-crosslinked species detected in 32D. E2/E3 cells were displaced by HRG and betacellulin (BTC) but not by other EGF-like ligands that were analyzed. EGF, BTC and HRG also induced receptor tyrosine phosphorylation, activation of downstream signaling molecules and proliferation of 32D.E2/E3 cells. 32D transfectants were also generated which expressed an ErbB3-EGFR chimera alone (32D.E3-E1) or in combination with ErbB2 (32D. E2/E3-E1). While HRG stimulation of 32D.E3-E1 cells resulted in DNA synthesis and receptor phosphorylation, EGF and BTC were inactive. However, EGF and BTC were as effective as HRG in mediating signaling when ErbB2 was co-expressed with the chimera in the 32D.E2/E3-E1 transfectant. These results provide evidence that ErbB2/ErbB3 binding sites for EGF and BTC are formed by a previously undescribed mechanism that requires co-expression of two distinct receptors. Additional data utilizing MDA MB134 human breast carcinoma cells, which naturally express ErbB2 and ErbB3 in the absence of EGFRs, supported the results obtained employing 32D cells and suggest that EGF and BTC may contribute to the progression of carcinomas that co-express ErbB2 and ErbB3.     

Serotonin(2) receptors mediate respiratory recovery after cervical spinal cord hemisection in adult rats.

The aim of the present study was to specifically investigate the involvement of serotonin [5-hydroxytryptamine (5-HT(2))] receptors in 5-HT-mediated respiratory recovery after cervical hemisection. Experiments were conducted on C(2) spinal cord-hemisected, anesthetized (chloral hydrate, 400 mg/kg ip), vagotomized, pancuronium- paralyzed, and artificially ventilated female Sprague-Dawley rats in which CO(2) levels were monitored and maintained. Twenty-four hours after spinal hemisection, the ipsilateral phrenic nerve displayed no respiratory-related activity indicative of a functionally complete hemisection. Intravenous administration of the 5-HT(2A/2C)-receptor agonist (+/-)-2,5-dimethoxy-4-iodoamphetamine hydrochloride (DOI) induced respiratory-related activity in the phrenic nerve ipsilateral to hemisection under conditions in which CO(2) was maintained at constant levels and augmented the activity induced under conditions of hypercapnia. The effects of DOI were found to be dose dependent, and the recovery of activity could be maintained for up to 2 h after a single injection. DOI-induced recovery was attenuated by the 5-HT(2)-receptor antagonist ketanserin but not with the 5-HT(2C)-receptor antagonist RS-102221, suggesting that 5-HT(2A) and not necessarily 5-HT(2C) receptors may be involved in the induction of respiratory recovery after cervical spinal cord injury.     

Muscarinic acetylcholine receptors mediate oligodendrocyte progenitor survival through Src-like tyrosine kinases and PI3K/Akt pathways

The function of muscarinic acetylcholine receptors expressed in oligodendrocytes and in myelin has remained largely undetermined. Here we present evidence that incubation of oligodendrocyte progenitors, deprived of growth factor, with the acetylcholine analog carbachol significantly reduced cell death by apoptosis and blocked caspase-3 cleavage. This protective effect was reversed by atropine, a muscarinic acetylcholine receptor antagonist, as well as by specific inhibitors of intracellular signaling molecules, including phosphatidylinositol 3-kinase (Wortmannin and LY294002), Akt (Akt inhibitor III) and Src-like tyrosine kinases (PP2), but not by the mitogen-activated protein kinase kinase inhibitor, PD98059. Activation of Akt by carbachol was antagonized by atropine and inhibited by LY294002 and PP2. The Src-like tyrosine kinase inhibitor, PP2, also reduced carbachol stimulation of extracellular signal-regulated kinases 1/2 and cAMP-response element binding protein in a dose-dependent manner. Furthermore, carbachol increased tyrosine-phosphorylation of Fyn, a member of the Src-like tyrosine kinases. These results indicate that muscarinic acetylcholine receptors play an important role in oligodendrocyte progenitor survival through transduction pathways involving activation of Src-like tyrosine kinases and phosphatidylinositol 3-kinase/Akt.     

大鼠杏仁核5-HT_3受体参与免疫调制

实验通过大鼠侧脑室和杏仁核给予 5 HT3受体激动剂 1 phenylbiguanide (PBG) ,用 3H TdR掺入法测定脾细胞丝裂原 (concanavalinA ,ConA和lipopolysaccharide ,LPS)刺激增殖效应 ,用活化脾细胞增殖法测定IL 2生成 ,MTT法测定自然杀伤 (naturalkiller,NK)细胞活性和用放射免疫测定血浆皮质酮水平 ,以探讨大鼠杏仁核 5 HT3受体在免疫调控中的作用。结果表明 :5 HT3受体拮抗剂granisetron (GNT ,0 1～ 0 4mg/kgip)剂量依赖地增强ConA和LPS刺激的脾细胞增殖 ,作用在连续给药 5d最明显 ;双侧脑室给予PBG ( 5 μg/side)可增强ConA和LPS刺激的脾细胞增殖效应 ,作用在连续给药 3d最明显 ;双侧和单侧中央杏仁核给予PBG 0 5 μg均增强ConA刺激的脾细胞增殖和IL 2生成 ;底内侧杏仁核给予同剂量PBG仅增强LPS刺激的脾细胞增殖效应 ,不影响ConA刺激的脾细胞增殖和IL 2生成 ;中央杏仁核给予PBG升高血浆皮质酮的作用较底内侧杏仁核给予等量PBG引起的升高血浆皮质酮作用明显 (P <0 0 1)。侧脑室、中央杏仁核和底内杏仁核给予PBG对丝裂原刺激的脾细胞增殖效应影响不同 ,但均被同时同部位给予GNT所拮抗 ,提示杏仁核中央核和底内侧核的 5 HT3受体可能以不同方式参与ConA或LPS刺激的脾细胞增殖效应的调制     

大鼠杏仁核5—HT3受体参与免疫调制

实验通过大鼠侧脑室和杏仁核给予5－HT3受体激动剂1－phenylbiguanide(PBG),用3H－TdR掺入法测定脾细胞丝裂原（concanavalin A,Con A和lipopolysaccharide,LPS)刺激增殖效应,用活化脾细胞增殖法测定IL－2生成,MTT法测定自然杀伤（natural killer,NK)细胞活性和用放射免疫测定血浆皮质酮水平,以探讨大鼠杏仁核5－HT3受体在免疫调控中的作用。结果表明：5－HT3受体拮抗剂granisetron(GNT,0.1-0.4mg/kg ip)剂量依赖地增强Con A和LPS刺激的脾细胞增殖,作用在连续给药5d最明显,双侧脑室给予PBG（5ug/side)可增强ConA和LPS刺激的脾细胞增殖效应,作用在连续给药3d最明显,双侧和单侧中共杏仁核给予PBG0．5ug均增强ConA刺激的脾细胞增殖和IL－2生成,底内侧杏仁核给予同剂量PBG仅增强LPS刺激的脾细胞增殖效应,不影响ConA刺激的脾细胞增殖和IL－2生成,中央杏仁核给予PBG升高血浆皮质酮的作用较底侧杏仁核给予等量PBG引起的升高血浆皮质酮作用明显（P＜0．01）,侧脑室,中央杏仁核和底内杏仁核给予PBG对丝裂原刺激的脾细胞增殖效应影响不同,但均被同时同部位给予GNT所拮抗,提示杏仁核中央核和底内侧核的5－HT3受体可能以不同方式参与ConA或LPS刺激的脾细胞增殖效应的调制。     

Serotonin-type 3 receptors mediate intestinal Polycose- and glucose-induced suppression of intake

Ondansetron, a selective serotonin-type 3 (5-HT(3)) receptor antagonist, was used to test the hypothesis that duodenal infusion of isosmotic solutions of Polycose or its hydrolytic product glucose suppressed intake through 5-HT(3) receptors. Polycose suppressed sucrose intake across both concentrations infused (132 mM, 7.6 +/- 0.6 ml; 263 mM, 2.3 +/- 0.5 ml), compared with intake under control conditions (12.6 +/- 0.3 ml, P <0.001). Pretreatment with 1.0 mg/kg ondansetron attenuated reduction of sucrose intake induced only by the highest concentration of Polycose (4.6 +/- 0.8 ml, P = 0.004). Dose-response testing revealed that suppression of food intake by 263 mM Polycose was equally attenuated by ondansetron administered at 1.0, 2.0, and 5.0 mg/kg but not when given at 0.125, 0.25, and 0.5 mg/kg. Acarbose, an alpha-glucosidase inhibitor, attenuated Polycose-induced suppression of food intake, and pretreatment with 1.0 mg/kg ondansetron had no further effect. Suppression of intake after 990 mM glucose but not mannitol infusion was attenuated by pretreatment with 1.0 mg/kg ondansetron. The competitive SGLT(1) inhibitor, phloridzin, had no effect on 60-min 990 mM glucose-induced suppression of intake or the ability of ondansetron to attenuate this suppression of intake. Conversely, glucose-induced suppression of intake was attenuated by phloridzin at earlier time points and further attenuated when rats were pretreated with 1.0 mg/kg ondansetron. Ondansetron administration alone had no effect on intake at any dose tested. We conclude that 5-HT(3) receptors participate in the inhibition of food intake by intraduodenal infusion of carbohydrate solutions through a posthydrolytic, preabsorptive mechanism.     

Serotonin 5-HT(3) receptors on mechanosensitive neurons with cardiac afferents

In rats, the mechanosensitive cardiorenal baroreflex influencing renal excretory function might be impaired by serotonin occurring in coronary arteries, e.g., in hypertension. Because the afferent limb of this reflex could be affected, we investigated the responses of nodose ganglion cells (one neuron of reflex) to osmotic, mechanical stress in presence or absence of the serotonin 5-HT(3) receptor agonist phenylbiguanide (PBG). Current-voltage relationships (from -100 to +50 mV) were obtained using cell patch recordings while the cells were exposed to control or hypoosmotic solutions to induce mechanical stress. This protocol was repeated after low doses of PBG (10 microM), angiotensin II (10 nM), or the stretch-activated channel blocker gadolinium (20 microM) were added to the extracellular medium (EM). Hypoosmotic EM induced significant changes in cellular conductance. The full-range current-voltage relationship allowed for the calculation of a mean reversal potential of -13 +/- 1.2 mV with respect to this change in cellular conductance (n = 44). This increase in conductance was impaired after addition of either PBG or gadolinium to the EM,which was statistically evaluated at a voltage of -80 mV, where influences of voltage-gated channels are not likely to interfere with the responses recorded. The serotonin 5-HT(3) receptor antagonist tropisetron (10 nM) prevented the PBG effect on conductance responses. Angiotensin II had no influence. Hence, serotonin might decrease the mechanical sensitivity of afferent cardiac nerves controlling renal sympathetic nerve activity.     

Serotonin stimulates endotoxin translocation via 5-HT3 receptors in the rat ileum

Because few previous studies have investigated the mechanisms of endotoxin translocation induced by intestinal obstruction, we aimed to clarify whether or not serotonin [5-hydroxytryptamine (5-HT)], which is released from enterochromaffin (EC) cells, is responsible for alterations of the mucosal permeability to endotoxin and to identify the 5-HT receptor subtypes that mediate this action. FITC-labeled LPS (FITC-LPS) was injected into the ileum of rats, and the FITC-LPS level in the superior mesenteric vein was subsequently measured by using a fluorescence spectrophotometer. To measure the 5-HT release induced by high intraluminal pressure, ex vivo preparation of vascularly and luminally perfused rat ileum was used. Results demonstrated that elevated intraluminal pressure stimulates the translocation of FITC-LPS and the release of 5-HT from the EC cells into the intestinal lumen but not into the portal circulation. This FITC-LPS translocation, which was stimulated by exogenously applied 5-HT in the lumen and the jugular vein, was inhibited by 5-HT(3) receptor antagonist administration both intaluminally and intravenously. The stimulatory effect of elevated intraluminal pressure on the translocation of FITC-LPS was inhibited by the intraluminal and intravenous administration of 5-HT(3) receptor antagonist. These results suggest that 5-HT released from EC cells may be involved in the translocation of FITC-LPS induced by elevated intraluminal pressure and that this effect is mediated by 5-HT(3) receptors that may be located in the intestinal epithelium.     

Enhancement of the function of neutrophil type-1 and type-3 complement receptors by human leukocyte inhibitory factor (LIF)

The lymphokine leukocyte inhibitory factor (LIF) has previously been documented to enhance several neutrophil (PMN) functions, including stimulated chemotaxis and superoxide generation, phagocytosis and adherence of opsonized targets, and antibody-dependent cellular cytotoxicity. The present studies were designed to investigate the effects of LIF on PMN function mediated by the complement components C3b and C3bi. LIF induced a dose-dependent increase in superoxide production generated by opsonized zymosan (up to 97.1 +/- 31.4% at 16 U LIF/ml; P less than 0.01). While neither control nor LIF-treated PMN were capable of inducing phagocytosis of either C3b- or C3bi-opsonized sheep erythrocytes (E) directly, exposure to LIF caused a significant (P less than 0.05) increase in their adherence to E (137.4 and 59.4%, respectively). Specificity for complement receptor function was confirmed by the ability of anti-CR1 antibody to block adherence of LIF-treated PMN to EAC3b (77.0% inhibition) and anti-CR3 antibody to block adherence to EAC3bi (70.2% inhibition). Increased C3b and C3bi function may have been due, at least in part, to increased expression of their respective surface membrane receptors. Thus, using indirect immunofluorescence, LIF induced a 38.2% increase in fluorescence of the anti-CR1 antibody and a 96.1% increase in anti-CR3 binding. These studies describe an additional mechanism through which LIF may have an important pro-inflammatory role in vivo.     

P2X7 nucleotide receptors mediate blebbing in osteoblasts through a pathway involving lysophosphatidic acid

Extracellular nucleotides, released in response to mechanical or inflammatory stimuli, signal through P2 receptors in many cell types, including osteoblasts. P2X7 receptors are ATP-gated cation channels that can induce formation of large membrane pores. Disruption of the gene encoding the P2X7 receptor leads to decreased periosteal bone formation and insensitivity of the skeleton to mechanical stimulation. Our purpose was to investigate signaling pathways coupled to P2X7 activation in osteoblasts. Live cell imaging showed that ATP or 2 ',3 '-O-(4-benzoylbenzoyl)-ATP (BzATP), but not UTP, UDP, or 2-methylthio-ADP, induced dynamic membrane blebbing in calvarial osteoblasts. Blebbing was observed in calvarial cells from wildtype but not P2X7 knock-out mice. P2X7 receptors coupled to activation of phospholipase D and A2, inhibition of which suppressed BzATP-induced blebbing. Activation of these phospholipases leads to production of lysophosphatidic acid (LPA). LPA caused dynamic blebbing in osteoblasts from both wild-type and P2X7 knock-out mice, similar to that induced by BzATP in wildtype cells. However, LPA-induced blebbing was more rapid in onset and was not affected by inhibition of phospholipase D or A2. Blockade or desensitization of LPA receptors suppressed blebbing in response to LPA and BzATP, without affecting P2X7-stimulated pore formation. Thus, LPA functions downstream of P2X7 receptors to induce membrane blebbing. Furthermore, inhibition of Rho-associated kinase abolished blebbing induced by both BzATP and LPA. In summary, we propose a novel signaling axis that links P2X7 receptors through phospholipases to production of LPA and activation of Rho-associated kinase. This pathway may contribute to P2X7-stimulated osteogenesis during skeletal development and mechanotransduction.     

Muscarinic M2 receptors mediate transactivation of EGF receptor through Fyn kinase and without matrix metalloproteases

Transactivation of epidermal growth factor receptor (EGFR) by G protein-coupled receptors (GPCRs) has been attributed to the activation of matrix metalloproteases (MMPs) and the release of EGF family ligands such as HB-EGF. This mode of transactivation leads to signalling downstream of EGFR which is indistinguishable from that induced by the ligand. Here we provide evidence that in the COS-7 cell model EGFR transactivation via the muscarinic M2 receptor (M2R) is independent of MMPs and results in an incomplete EGFR signalling including ERK and Akt but not PLCgamma1. Using dominant-negative mutants of c-Src and Fyn and Src-deficient SYF cells as well as by co-immunoprecipitation studies, we can demonstrate that the M2R-mediated transactivation of EGFR specifically involves Fyn but not c-Src or Yes. This specific role of Fyn can be verified in SH-SY5Y human neuroblastoma cells with endogenously expressed M2 receptors.     

High-altitude hypoxia induces disorders of the brainendocrine-immune network through activation of corticotropin-releasing factor and its type-1 receptors

High-altitude hypoxia can induce physiological dysfunction and mountain sickness,but the underlying mechanism is not fully understood.Corticotrophin-releasing factor(CRF) and CRF type-1 receptors(CRFR1) are members of the CRF family and the essential controllers of the physiological activity of the hypothalamo-pituitary-adrenal(HPA) axis and modulators of endocrine and behavioral activity in response to various stressors.We have previously found that high-altitude hypoxia induces disorders of the brain-endocrine-immune network through activation of CRF and CRFR1 in the brain and periphery that include activation of the HPA axis in a time-and dose-dependent manner,impaired or improved learning and memory,and anxiety-like behavioral change.Meanwhile,hypoxia induces dysfunctions of the hypothalamo-pituitary-endocrine and immune systems,including suppression of growth and development,as well as inhibition of reproductive,metabolic and immune functions.In contrast,the small mammals that live on the Qinghai-Tibet Plateau alpine meadow display low responsiveness to extreme high-altitudehypoxia challenge,suggesting well-acclimatized genes and a physiological strategy that developed during evolution through interactions between the genes and environment.All the findings provide evidence for understanding the neuroendocrine mechanisms of hypoxia-induced physiological dysfunction.This review extends these findings.     

Cloned rat M3 muscarinic receptors mediate phosphoinositide hydrolysis but not adenylate cyclase inhibition

Rat M3 mAChR subtype was stably expressed in RAT 1 cells. Investigation of the pharmacological and biochemical properties of the cloned M3 receptors revealed that they mediate phosphoinositide hydrolysis but not adenylate cyclase inhibition. The similarities and differences between the properties of cloned rat M1 and M3 receptors are discussed.     

Targeting the inflammasome and adenosine type-3 receptors improves outcome of antibiotic therapy in murine anthrax

AIM: To establish whether activation of adenosine type-3 receptors (A3Rs) and inhibition of interleukin-1β-induced inflammation is beneficial in combination with antibiotic therapy to increase survival of mice challenged with anthrax spores.METHODS: DBA/2 mice were challenged with Bacillus anthracis spores of the toxigenic Sterne strain 43F2. Survival of animals was monitored for 15 d. Ciprofloxacin treatment (50 mg/kg, once daily, intraperitoneally) was initiated at day +1 simultaneously with the administration of inhibitors, and continued for 10 d. Two doses (2.5 mg/kg and 12.5 mg/kg) of acetyl-tyrosyl-valyl-alanyl-aspartyl-chloromethylketone (YVAD) and three doses (0.05, 0.15 and 0.3 mg/kg) of 1-[2-Chloro-6-[[(3-iodophenyl) methyl]amino]-9H-purin-9-yl]-1-deoxy-N-methyl-β-D- ribofuranuronamide (Cl-IB-MECA) were tested. Animals received YVAD on days 1-4, and Cl-IB-MECA on days 1-10 once daily, subcutaneously. Human lung epithelial cells in culture were challenged with spores or edema toxin and the effects of IB-MECA on phosphorylation of AKT and generation of cAMP were tested.RESULTS: We showed that the outcome of antibiotic treatment in a murine anthrax model could be substantially improved by co-administration of the caspase-1/4 inhibitor YVAD and the A3R agonist Cl-IB-MECA. Combination treatment with these substances and ciprofloxacin resulted in up to 90% synergistic protection. All untreated mice died, and antibiotic alone protected only 30% of animals. We conclude that both substances target the aberrant host signaling that underpins anthrax mortality.CONCLUSION: Our findings suggest new possibilities for combination therapy of anthrax with antibiotics, A3R agonists and caspase-1 inhibitors.     

E-prostanoid-3 receptors mediate the proinflammatory actions of prostaglandin E2 in acute cutaneous inflammation

PGs are derived from arachidonic acid by PG-endoperoxide synthase (PTGS)-1 and PTGS2. Although enhanced levels of PGs are present during acute and chronic inflammation, a functional role for prostanoids in inflammation has not been clearly defined. Using a series of genetically engineered mice, we find that PTGS1 has the capacity to induce acute inflammation, but PTGS2 has negligible effects on the initiation of this response. Furthermore, we show that the contribution of PTGS1 is mediated by PGE(2) acting through the E-prostanoid (EP)3 receptor. Moreover, in the absence of EP3 receptors, inflammation is markedly attenuated, and the addition of nonsteroidal anti-inflammatory agents does not further impair the response. These studies demonstrate that PGE(2) promotes acute inflammation by activating EP3 receptors and suggest that EP3 receptors may be useful targets for anti-inflammatory therapy.