IL-18 binding protein increases spontaneous and IL-1-induced prostaglandin production via inhibition of IFN-gamma

IL-18 shares with IL-1 the same family of receptors and several identical signal transduction pathways. Because of these similarities, IL-18 was investigated for its ability to induce prostaglandin E(2) (PGE(2)) synthesis in human peripheral blood mononuclear cells (PBMC), a prominent, proinflammatory property of IL-1. IL-18 was highly active in PBMC by inducing the synthesis of the chemokine IL-8; however, no induction of PGE(2) synthesis nor cyclooxygenase type-2 gene expression was observed in PBMC stimulated with IL-18. In the same cultures, IL-1beta induced a 12-fold increase in PGE(2). Although IL-1beta-induced IL-8 synthesis was augmented 3-fold by IL-18, IL-18 suppressed IL-1beta-induced PGE(2) production by 40%. The suppressive effect of IL-18 on PGE(2) production was mediated by interferon (IFN)-gamma because anti-human IFN-gamma-antibody prevented IL-18-induced reduction in PGE(2). Consistent with these observations, IL-12, a known inducer of IFN-gamma, augmented IL-1beta-induced IFN-gamma but suppressed IL-1beta-induced PGE(2) by 75%. IL-18 binding protein (IL-18BP) is a naturally occurring and specific inhibitor of IL-18. When recombinant IL-18BP was added to PBMC cultures, unexpectedly, spontaneous PGE(2) production increased. PGE(2) production was also increased by the addition of IL-18BP to PBMC stimulated with either IL-1beta or IL-12 and also in whole blood cultures stimulated with Staphylococcus epidermidis. These studies demonstrate that IL-18BP decreases endogenous IL-18 activity by reducing IFN-gamma-mediated responses.     

Differential effects of interleukin-15 (IL-15) and IL-2 on human neutrophils: modulation of phagocytosis, cytoskeleton rearrangement, gene expression, and apoptosis by IL-15

Human neutrophils have been shown recently to express both the beta and the gamma chains of the interleukin-2 receptor (IL-2R). IL-15, a cytokine that has recently been cloned and characterized, was found to share many of the biological functions of IL-2 and is known to mediate signals through IL-2R beta and IL-2R gamma. In recent studies, we observed that IL-2 exerts few effects on various neutrophil functions, but information on IL-15-neutrophil interactions is lacking. In this study, we observed that IL-15, in contrast to IL-2, induces important morphological cell shape changes that are typical of activated neutrophils. Furthermore, phagocytosis of opsonized sheep red blood cells was significantly increased by IL-15 but not by IL-2. However, similar to IL-2, IL-15 did not modulate the oxidative burst response. Furthermore, we observed that de novo RNA synthesis is increased in neutrophils by IL-15 along with de novo protein synthesis, whereas no significant effect of IL-2 was noted. Among the different proteins that were found to be upregulated by IL-15, one was identified by microsequencing as the cytoskeletal protein actin. Finally, we found that IL-15 delays apoptosis of neutrophils more efficiently than IL-2 when evaluated by both microscopic observations and flow cytometry procedures. Furthermore, this phenomenon was dose-dependent (10 to 500 ng/mL), and, at 500 ng/mL, IL-15 delayed apoptosis as strongly as granulocyte-macrophage colony-stimulating factor. This study is the first to show that IL-15 is a significant neutrophil agonist. Moreover, in view of the differential effects of IL-15 and IL-2 on this cell type, our results support the existence of a specific IL-15R component(s) on human neutrophils.     

Secretion of IFN-gamma and not IL-2 by anergic human T cells correlates with assembly of an immature immune synapse

We report differences in the supramolecular organization of the immunologic synapse (IS) formed by resting and anergic human T cells with agonist peptide-loaded antigen-presenting cells (APCs). T cells reactive to influenza A hemagglutinin peptide or Fel d 1 peptide 4 were rendered both anergic and regulatory by incubation with high doses of agonist peptide in the absence of APCs. At the IS between resting T cells and peptide-loaded APCs, both CD3epsilon and CD3zeta initially accumulate within a ring or arc before redistributing within 30 minutes to single or multiple foci more central to the contact. In contrast, at synapses formed by anergized T cells, CD3epsilon and CD3zeta remained organized within an arc or ring and failed to redistribute centrally. However, intercellular communication between anergic human T cells and agonist peptide-loaded APCs was not a null event, since it triggered secretion of T-cell interferon gamma (IFN-gamma) but not, for example, interleukin 2 (IL-2). Thus, distinct organizations of CD3 at the T-cell IS correlate with different cytokine profiles; the mature IS formed by resting T cells correlates with their production of both IFN-gamma and IL-2, whereas the immature IS formed by anergic T cells seems able to facilitate IFN-gamma but not IL-2 production.     

Inhibition of IL-1beta-dependent prostaglandin E2 release by antisense microsomal prostaglandin E synthase 1 oligonucleotides in A549 cells

The metabolism of arachidonic acid through the cyclooxygenase pathway is a highly regulated cellular process that results in the formation of PGH2. This unstable intermediate can be enzymatically metabolized to PGE2 by the actions of a microsomal 17 kDa PGE synthase (mPGES1). Treatment of A549 cells with IL-1beta for 24 h resulted in a twofold increase in mPGES1 mRNA, protein expression, and PGES specific activity. To understand the relationship between expression of mPGES1 and PGE2 formation, IL-1beta treated cells were incubated with increasing concentrations of antisense oligonucleotides (ASO) and their effects compared to cells treated with reverse sense oligonucleotides (RSO) designed against the ATG translation initiation codon of mPGES1. Incubation with ASO resulted in a 44% reduction in mRNA expression level as compared to RSO-treated cells. Microsomal preparations isolated from ASO- and RSO-treated cells were analyzed for their ability to convert PGH2 to PGE2 in the presence 2.5 mM reduced glutathione. An approximate 50% reduction (ASO: 1.8 nmol/min/mg, RSO: 3.7 nmol/min/mg) in PGES activity, protein expression by immunodetection, and extracellular PGE2 release was detected in these samples. As a control in these studies, the protein levels of COX2 and secreted IL-8 were quantified; no change in these levels was observed. These results demonstrate the direct association between mPGES1 expression, its enzymatic activity, and total PGE2 production following an inflammatory stimulus.     

Evaluation of In Vitro Production of IFN-gamma, IL-10, IL-12 and IL-13 by Blood Cells in Patients with Cutaneous Leishmaniasis Lesions

This study investigated the in vitro production of interferon-gamma, interleukin (IL)-10, IL-12, and IL-13, after antigenic stimulation of the cells (with Leishmania antigen and lipopolysaccharide) using whole blood from patients with cutaneous leishmaniasis lesions caused by Leishmania tropica and in normal volunteers with history of cutaneous leishmaniasis.ELISA results showed that the mean production of interferon-gamma by cells of whole blood in patients with lesions in response to Leishmania antigen was significantly lower than corresponding values in volunteers with history of cutaneous leishmaniasis (P< 0.05) and significantly higher levels of IL-10 production in patients with lesions were observed compared with cured volunteers of the disease (P<0.01). A similar level of IL-12, including p40 subunit of IL-12, was detected in both groups tested in this study in response to stimulation of parasite antigen. The levels of the IL-13 after stimulation with Leishmania antigen were significantly more in patients compared with volunteers with history of cutaneous leishmaniasis (P< 0.01). There was no significant difference in the mean production of IFN-gamma, IL-10, IL-12 and IL-13 by PHA or LPS stimulated cells from patients with lesions and volunteers with history of the disease, indicating that there was no qualitative defect in cytokine production in these patients.In this study, we have detected the decreased production of interferon- gamma by cells of patients with lesions of cutaneous leishmaniasis in response to parasite antigen and unbalanced production of regulatory cytokines such as IL-10 and IL-13 using the whole-blood stimulation assay technique. The required small volume of blood and the rapid set up time are the advantages in this assay technique. Using this assay for further immunodetection of cytokines may confirm its value for clinical investigation.     

Enhancement of the host immune responses in cutaneous T-cell lymphoma by CpG oligodeoxynucleotides and IL-15

Patients with advanced cutaneous T-cell lymphoma (CTCL) exhibit profound defects in cell-mediated immunity. Host immune functions appear to play an integral role in mediating disease-controlling responses in CTCL, therefore we investigated the effects of synthetic oligode-oxynucleotides with CpG motifs (CpG ODN), which have been recognized as immune stimulatory by virtue of activation of dendritic cells (DCs) following binding to Toll-like receptor (TLR) 9. Peripheral blood mononuclear cells (PBMCs) from patients with advanced CTCL (erythroderma with circulating malignant T cells) and healthy volunteers were cultured with either CpG-A or CpG-B ODN. Patients' PBMCs exhibited marked induction of interferon-alpha (IFN-alpha) release following culture with CpG-A. Similarly significant activation of NK cells and CD8 T cells occurred as assessed by up-modulation of CD69 expression and by natural killer lytic activity. Nevertheless, the PBMCs of patients exhibited blunted responses to CpG-A compared to healthy volunteers. In such cases, IL-15 was capable of producing levels of NK activation that were superior to CpG-A, while the combined effects of CpG-A plus IL-15 induced maximal activation of NK cells and further enhanced activation of CD8 T cells. These findings have important implications for the potential enhancement of antitumor immunity among patients with advanced CTCL.     

IL-18, although antiallergic when administered with IL-12, stimulates IL-4 and histamine release by basophils

IL-18 is a proinflammatory cytokine that plays an important role in natural killer cell activation and T helper 1 (Th1) cell responses. Mast cells and basophils are major inducers and effectors of allergic inflammation. Here we show that basophils and mast cells derived by culture of bone marrow cells with IL-3 for 10 days express IL-18Ralpha chain and that basophils produce large amounts of IL-4 and IL-13 in response to stimulation with IL-3 and IL-18. Injection of IL-12 and IL-18 inhibits IgE production in helminth-infected wild-type mice and abolishes the capacity of their basophils to produce IL-4 and IL-13 in response to stimulation either with IL-3 and IL-18 or with FcepsilonR cross-linkage. By contrast, this combination of cytokines actually increases IgE levels in helminth-infected IFN-gamma(-/-) mice and enhances IL-4 and IL-13 production by their basophils. Furthermore, injection of IL-18 alone enhances basophil production of IL-4 and histamine both in wild-type and IFN-gamma(-/-) mice. Thus, IL-18 has the potential to stimulate basophils but, when given with IL-12, exhibits an antiallergic action in vivo.     

Differential regulation of T-cell growth by IL-2 and IL-15

Although interleukin 2 (IL-2) and IL-15 signal through the common gamma chain (gammac) and through IL-2 receptor beta-chain (CD122) subunits, they direct distinct physiologic and immunotherapeutic responses in T cells. The present study provides some insight into why IL-2 and IL-15 differentially regulate T-cell function by revealing that these cytokines are strikingly distinct in their ability to control protein synthesis and T-cell mass. IL-2 and IL-15 are shown to be equivalent mitogens for antigen-stimulated CD8(+) T cells but not for equivalent growth factors. Antigen-primed T cells cannot autonomously maintain amino acid incorporation or de novo protein synthesis without exogenous cytokine stimulation. Both IL-2 and IL-15 induce amino acid uptake and protein synthesis in antigen-activated T cells; however, the IL-2 response is strikingly more potent than the IL-15 response. The differential action of IL-2 and IL-15 on amino acid uptake and protein synthesis is explained by temporal differences in signaling induced by these 2 cytokines. Hence, the present results show that cytokines that are equivalent mitogens can have different potency in terms of regulating protein synthesis and cell growth.     

Role of caspase-1 in nuclear translocation of IL-37, release of the cytokine,and IL-37 inhibition of innate immune responses

IL-37 is a fundamental inhibitor of innate immunity. Human IL-37 has a caspase-1 cleavage site and translocates to the nucleus upon LPS stimulation. Here, we investigated whether caspase-1 processing affects IL-37–mediated suppression of LPS-induced cytokines and the release from cells by analyzing a caspase-1 cleavage site mutant IL-37 (IL-37D20A). Nuclear translocation of IL-37D20A is significantly impaired compared with WT IL-37 in transfected cells. LPS-induced IL-6 was decreased in cells expressing WT IL-37 but not IL-37D20A. The function of IL-37 in transfected bone marrow-derived macrophages is nucleotide-binding oligomerization domain-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome-dependent, because IL-37 transfection in apoptosis-associated speck-like protein containing a carboxyl-terminal caspase recruitment domain- and NLRP3-deficient cells does not reduce levels of IL-6 and IL-1β upon LPS stimulation. IL-37–expressing macrophages release both precursor and mature IL-37, but only the externalization of mature IL-37 was dependent on ATP. Precursor and mature IL-37 was also secreted from human dendritic cells and peripheral blood mononuclear cells. To determine whether IL-37 is active in the extracellular compartment, we pretreated IL-37 transgenic mice with IL-37–neutralizing antibodies before LPS challenge. In IL-37–expressing mice, neutralizing IL-37 antibodies reversed the suppression of LPS-induced serum IL-6. In contrast, the addition of neutralizing antibody did not reverse suppression of LPS-induced IL-6 in mouse macrophages transfected with IL-37. Although caspase-1 is required for nuclear translocation of intracellular IL-37 and for secretion of mature IL-37, the release of the IL-37 precursor is independent of caspase-1 activation. IL-37 now emerges as a dual-function cytokine with intra- and extracellular properties for suppressing innate inflammation.With the exception of the IL-1 receptor antagonist, members of the IL-1 family are first synthesized as precursor molecules containing a propeptide domain lacking a classical signal sequence (1). Caspase-1 has emerged as the main intracellular processing enzyme responsible for maturation of active IL-1β and IL-18, which are then released into the extracellular space, as shown for IL-1β and IL-18 (2, 3). The IL-1 family member IL-37 is also synthesized as a precursor and is processed to its mature form upon LPS treatment (4, 5). Caspase-1 seems to be the main enzyme responsible for the in vitro maturation of IL-37 in comparison to caspase-4 and granzyme B (4). A putative cleavage site for caspase-1 is located in exon 1 between residues D20 and E21 of IL-37 (4). HEK 293 or CHO cells transfected with the IL-37 precursor release IL-37 starting at amino acid V46, suggesting a second cleavage site in the sequence encoded by exon 2 (6). We previously demonstrated that processing of IL-37 is only partially inhibited by caspase-1 inhibitors, indicating that caspase-1 is not the only enzyme responsible for the processing of IL-37 (5).In our previous study, we showed that transgenic expression of human IL-37 in a mouse macrophage line significantly suppressed the production of proinflammatory cytokines and chemokines (5). Furthermore, we reported that IL-37 has significant anti-inflammatory effects in an in vivo model of septic shock and dextran sulfate sodium salt-induced colitis (7, 8). Here, we investigate the role of caspase-1 processing on the cytokine-suppressing function of IL-37. We introduced a point mutation into the caspase-1 cleavage site in the IL-37 gene by site-directed mutagenesis and expressed mutant IL-37 in RAW264.7 (RAW) mouse macrophages. In addition, we studied the release of IL-37 from human peripheral blood mononuclear cells (PBMCs) and dendritic cells (DCs). The data indicate that the precursor and mature forms of IL-37 are secreted from activated cells upon inflammasome activation and that caspase-1 processing of IL-37 is important for its anti-inflammatory activity in vitro and in vivo.     

IL-7 and IL-15 differentially regulate CD8+ T-cell subsets during contraction of the immune response

Although it is known that interleukin-7 (IL-7) and IL-15 influence the survival and turnover of CD8+ T cells, less is known about how these cytokines affect different subsets during the course of the immune response. We find that IL-7 and IL-15 differentially regulate CD8+ T-cell subsets defined by KLRG1 and CD127 expression during the contraction phase of the immune response. The provision of IL-15, or the related cytokine IL-2, during contraction led to the preferential accumulation of KLRG1(hi)CD127(lo) CD8+ T cells, whereas provision of IL-7 instead favored the accumulation of KLRG1(lo)CD127(hi) cells. While IL-7 and IL-15 both induced proliferation of KLRG1(lo) cells, KLRG1(hi) cells exhibited an extraordinarily high level of resistance to cytokine-driven proliferation in vivo despite their dramatic accumulation upon IL-15 administration. These results suggest that IL-15 and IL-2 greatly improve the survival of KLRG1(hi) CD8+ T cells, which are usually destined to perish during contraction, without inducing proliferation. As the availability of IL-15 and IL-2 is enhanced during periods of extended inflammation, our results suggest a mechanism in which a population of cytokine-dependent KLRG1(hi) CD8+ T cells is temporarily retained for improved immunity. Consideration of these findings may aid in the development of immunotherapeutic strategies against infectious disease and cancer.     

Interleukin-21 receptor (IL-21R) is up-regulated by CD40 triggering and mediates proapoptotic signals in chronic lymphocytic leukemia B cells

Interleukin-21 (IL-21) is a member of the IL-2 cytokine family, which mediates proliferation or growth arrest and apoptosis of normal B cells, depending on their activation state. Here we demonstrate that surface IL-21 receptor (R) is expressed at variable levels by chronic lymphocytic leukemia (CLL) B cells freshly isolated from 33 different patients. IL-21R expression was up-regulated following cell stimulation via surface CD40. Therefore, IL-21 effects were more evident in CD40-activated CLL B cells. IL-21 induced an early signaling cascade in CLL B cells, which included JAK-1 and JAK-3 autophosphorylation and tyrosine phosphorylation of STAT-1, STAT-3, and STAT-5. IL-21 signaling failed to stimulate CLL B-cell proliferation, but induced their apoptosis. In addition, IL-21 counteracted the proliferative and antiapoptotic signals delivered by IL-15 to CLL B cells. IL-21-mediated apoptosis involved activation of caspase-8 and caspase-3, cleavage of Bid to its active form t-Bid, and cleavage of PARP and of p27Kip-1. Recent data indicate that CLL B cells require interaction with the microenvironment for their survival and expansion. The present findings thus provide a set of new mechanisms involved in the balance between cell-survival and apoptotic signals in CLL B cells.     

Role of IL-1 receptor-associated kinase-M (IRAK-M) in priming of immune and inflammatory responses by nitrogen bisphosphonates

Nitrogen bisphosphonates (NBPs) are commonly prescribed for osteoporosis but have also been found to induce inflammatory reactions and to delay the progression of breast cancer. The inflammatory and anticancer effects of the NBPs might be associated with an ability to modulate innate immune signaling. In mice, intraperitoneal NBP administration causes a rapid influx of neutrophils and monocytes that is dependent on the myeloid differentiation primary response gene 88 (MyD88) mediator of Toll-like receptor (TLR) and IL-1 signaling. Bone marrow chimeras demonstrate that this inflammatory response is partially dependent on TLR4 expression by hematopoietic cells and the IL-1 receptor on radioresistant cells. In vitro, NBPs directly stimulate neither murine bone marrow-derived mononuclear cells nor human peripheral blood mononuclear cells, but rather prime them to produce increased amounts of cytokines when exposed to IL-1 or TLR ligands. This potentiation is mediated by a reduction in IL-1 receptor-associated kinase-M, a negative regulator of MyD88-dependent signaling. In vivo, this property renders the NBPs as effective adjuvants that enhance both cellular and antibody responses to antigens.     

IL-15表达质粒联合HPV基因疫苗诱导细胞免疫应答的实验研究

目的研究白细胞介素15(IL-15)真核表达质粒,对人乳头瘤病毒(HPV)16E7基因疫苗所诱导的小鼠特异性细胞免疫应答的影响。方法构建含IL-15的真核表达质粒pcDNA3.1-IL-15。将该质粒与HPV16 E7基因疫苗通过肌肉注射方式免疫雌性BALB/c小鼠。基因免疫后测定其血清γ-干扰素(IFN-γ)水平;并制备脾淋巴细胞悬液,经体外E7蛋白再刺激后用MTT法检测其T淋巴细胞增殖情况。结果pcDNA3.1-IL-15与pcD-NA3.1-E7共同注射,可以提高免疫小鼠血清中IFN-γ水平至414.1pg/ml,与E7 空质粒组、E7组比较差异有统计学意义(P<0.01)。pcDNA3-1-IL-15与pcDNA3.1-E7共同注射,可以增强特异性T细胞增殖反应,OD570差值为1.313,与其他各组比较差异均有统计学意义(P<0.01)。结论IL-15真核表达质粒可以提高HPV16 E7基因疫苗的免疫原性,增强小鼠细胞免疫应答。     

Inhibition by amlodipine of activity evoked in isolated human coronary arteries by endothelin, prostaglandin F2 alpha and depolarization

Amlodipine, a dihydropyridine calcium antagonist has been examined on the rhythmic activity of isolated human coronary arteries. Amlodipine inhibited both the spontaneous rhythmic activity and the rhythmic activity evoked by prostaglandin F2 alpha and endothelin in isolated human coronary arteries. It also inhibited the contraction evoked by potassium depolarization. The action of amlodipine was characterized by slow onset and voltage dependency.     

Distinct cell types control lymphoid subset development by means of IL-15 and IL-15 receptor alpha expression

IL-15 and the IL-15 receptor (IL-15R)alpha chain are essential for normal development of naive CD8 T cells, intestinal intraepithelial lymphocytes (IEL), and natural killer (NK)/NK/T cells. However, whether IL-15R alpha expression by these subsets is necessary for their production and which cell type needs to produce IL-15 to drive development are unknown. We analyzed the requirements for IL-15 and IL-15R alpha expression by bone marrow-derived or parenchymal cells for mediating lymphocyte subset development. Naive CD8 T cell development required IL-15R alpha expression by both bone marrow-derived and parenchymal cells, whereas memory-phenotype CD8 T cells required IL-15R alpha expression only by hematopoietic cells. In contrast and surprisingly, the development of IEL subsets, particularly CD8 alpha alpha Thy1(-)V gamma 5(+) T cell antigen receptor gamma delta and the CD8 alpha alpha Thy1(-) T cell antigen receptor alpha beta IEL populations, depended completely on parenchymal cell expression of IL-15R alpha and IL-15 but not IL-15R beta. In the case of NK and NK/T cell generation and maturation, expression of IL-15 and IL-15R alpha by both parenchymal and hematopoietic cells was important, although the latter played the greatest role. These results demonstrated dichotomous mechanisms by which IL-15 regulated lymphoid development, interacting with distinct cell types depending on the developmental pathway.     

支气管哮喘在卡介菌多糖核酸治疗后IFN-γ、IL-4、IL-5的变化

目的　探讨支气管哮喘患者应用卡介菌多糖核酸 (PNBCG)注射液治疗之后 IFN-γ、IL - 4、IL - 5的血清值的变化。方法　 110例哮喘患者被随机分为卡介菌多糖核酸组、糖皮质激素治疗组和常规治疗组。卡组患者注射PNBCG注射液 0 .5 mg/ d× 90 d。结果　卡组治疗后 IFN-γ的血清值明显增高 ,是治疗前的 1.919倍 (95 % CI:2 82 .19;4 8.72 ) (p<0 .0 1)。 IL - 4、IL - 5的血清值明显降低 ,分别是治疗前的 0 .4 4倍 (95 % CI:1.4 2 ;0 .31) (p<0 .0 1)和0 .5 3倍 (95 % CI:2 1.12 ;- 4 .71) (p<0 .0 1)。结论　支气管哮喘患者在用 PNBCG后 IFN-γ的血清值升高 ,IL - 4、IL -5的血清值降低可能是哮喘患者病情转轻的原因     

人工肝支持系统治疗对重型肝炎患者血清TNF-α、IL-2、IL-10和IL-15水平的影响

目的 探讨重型肝炎患者人工肝支持系统(ALSS)治疗前后外周血中肿瘤坏死因子-α(TNF-α)、白细胞介素-2(IL-2)、白细胞介素-10(IL-10)、白细胞介素-15(IL-15)水平的变化。方法 应用血浆置换技术治疗重型肝炎患者59例，采用ELISA法于每次治疗前后检测TNF-α、IL-2、IL-10、IL-15水平，并观察其动态变化。结果 重型肝炎患者血清TNF-α水平明显高于对照组，IL-2水平明显低于对照组，ALSS治疗后TNF-α水平明显下降，而IL-2、IL-l0则呈上升趋势，治疗前后IL-15水平无明显变化。结论 ALSS治疗能降低重型肝炎患者TNF-α含量，并升高IN-2和IL-10水平，从而抑制炎性介质的产生，减轻免疫反应对肝细胞的损伤，提高重型肝炎的存活率。     

Neuronal adenosine release, and not astrocytic ATP release, mediates feedback inhibition of excitatory activity

Adenosine is a potent anticonvulsant acting on excitatory synapses through A1 receptors. Cellular release of ATP, and its subsequent extracellular enzymatic degradation to adenosine, could provide a powerful mechanism for astrocytes to control the activity of neural networks during high-intensity activity. Despite adenosine's importance, the cellular source of adenosine remains unclear. We report here that multiple enzymes degrade extracellular ATP in brain tissue, whereas only Nt5e degrades AMP to adenosine. However, endogenous A1 receptor activation during cortical seizures in vivo or heterosynaptic depression in situ is independent of Nt5e activity, and activation of astrocytic ATP release via Ca(2+) photolysis does not trigger synaptic depression. In contrast, selective activation of postsynaptic CA1 neurons leads to release of adenosine and synaptic depression. This study shows that adenosine-mediated synaptic depression is not a consequence of astrocytic ATP release, but is instead an autonomic feedback mechanism that suppresses excitatory transmission during prolonged activity.